Non-human animal with humanized gene, construction method therefor, and use thereof

Abstract

Provided are a non-human animal with humanized IL22, IL22RA1, IL10RB, IL20RA, and / or IL20RB genes, or a genome thereof, and use thereof in the field of biomedicine. All or part of the endogenous IL22, IL22RA1, IL10RB, IL20RA, and / or IL20RB genes of the non-human animal are replaced by corresponding human regions to obtain a modified non-human animal genome. The non-human animal comprising the modified non-human animal genome can normally express human or humanized IL22, IL22RA1, IL10RB, IL20RA, and / or IL20RB proteins, and can be used as an animal model for studying the signal mechanism of IL22, IL22RA1, IL10RB, IL20RA, and / or IL20RB and screening drugs for related diseases, demonstrating application value for the research and development of new drugs targeting immune targets.

Description

A genetically humanized non-human animal, its construction method and application Technical Field

[0001] This invention belongs to the field of animal genetic engineering and gene modification. Specifically, it relates to a non-human animal with humanized IL22, IL22RA1, IL10RB, IL20RA and / or IL20RB genes, its construction method, and its application in the biomedical field. Background Technology

[0002] The specific binding of cytokines to their receptors is a core mechanism for regulating immune responses, inflammatory responses, hematopoietic function, and tissue repair. This interaction not only serves as a diagnostic biomarker for early screening and prognostic assessment, but also plays a crucial role in treating infections, autoimmune diseases, cancer, and graft-versus-host disease by activating or blocking specific signaling pathways. Furthermore, it plays a key role in stem cell expansion and directed differentiation, vaccine adjuvant development, and immunomodulatory drug design.

[0003] The IL-20 subfamily includes cytokines IL-19, IL-20, IL-22, IL-24, and IL-26. The functions of the IL-20 subfamily include inducing innate defense mechanisms of epithelial cells against extracellular pathogens (such as bacteria and yeast), preventing tissue damage caused by infection and inflammation, stimulating epithelial cell proliferation, anti-apoptotic responses, and tissue remodeling and healing. Furthermore, its expression is elevated in autoimmune diseases such as psoriasis, rheumatoid arthritis (RA), and atopic dermatitis.

[0004] The IL-20 subfamily of cytokines has four receptors: IL22RA1, IL10RB, IL20RA, and IL20RB. These four receptors form dimers in pairs, which bind to IL-20 subfamily cytokines. Specifically:

[0005] The dimer formed by IL20RA and IL20RB is the receptor for IL19, IL20 and IL24; the dimer formed by IL20RA and IL10RB is the receptor for IL26; the dimer formed by IL22RA1 and IL10RB is the receptor for IL22; and the dimer formed by IL22RA1 and IL20RB is the receptor for IL20 and IL24.

[0006] In addition, IL10RB can also form a dimer with IL10RA and bind to IL10.

[0007] IL-20 family cytokines, upon binding to their receptors, can activate the downstream JAK / STAT signaling pathway. Phosphorylated STAT3 can cross into the nucleus and regulate the expression of downstream genes, thereby playing a role in immunomodulation.

[0008] With the continuous development and maturation of genetic engineering technology, replacing or substituting homologous genes in animals with human genes has become a reality. Developing humanized experimental animal models in this way represents the future direction of animal model development. Among these, gene-humanized animal models, which utilize gene editing technology to replace homologous genes in the animal genome with normal or mutated human genes, can establish normal or mutated gene animal models that more closely resemble human physiological or disease characteristics. Gene-humanized animals not only have significant application value—for example, they can improve and enhance cell or tissue transplantation models—but more importantly, due to the insertion of human gene fragments, the animals can express or partially express human proteins. These proteins can serve as targets for drugs that can only recognize the amino acid sequences of human proteins, making it possible to screen anti-human antibodies and other drugs at the animal level. However, due to the physiological and pathological differences between animals and humans, coupled with the complexity of genes (i.e. hereditary factors), the biggest challenge remains how to construct "effective" humanized animal models for new drug development (Scheer N, Snaith M, Wolf CR, Seibler J. Generation and utility of genetically humanized mouse models, Drug Discov Today; 18(23-24):1200-11, 2013).

[0009] Given the extensive involvement of IL20 subfamily cytokines and their receptors in the pathogenesis of various diseases, including immune diseases, and the enormous application value of targeting this signaling pathway, there is still an urgent need in the field to develop non-human animal models related to humanized signaling pathways of IL20 subfamily cytokines and their receptors in order to make preclinical trials more effective and minimize research and development failures. Summary of the Invention

[0010] In a first aspect, the present invention provides a non-human animal genome, wherein the non-human animal or the non-human animal genome comprises a genetically modified endogenous IL22RA1 gene encoding a modified IL22RA1 protein, and the non-human animal expresses the modified IL22RA1 protein.

[0011] Preferably, in the non-human animal or non-human animal genome, the nucleotide sequence encoding the corresponding segment of non-human animal endogenous IL22RA1 is replaced with the nucleotide sequence encoding the human IL22RA1 segment.

[0012] Preferably, the modified IL22RA1 protein comprises a cytoplasmic domain, a transmembrane domain, and an extracellular domain.

[0013] Preferably, the modified IL22RA1 protein contains an extracellular domain that is substantially the same as that of the human IL22RA1 protein.

[0014] Preferably, all or part of the extracellular domain comprises the amino acid sequence shown at positions 16-228 or 17-216 of SEQ ID NO: 27; or, comprises an amino acid sequence with an identity of at least 77%, 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown at positions 16-228 or 17-216 of SEQ ID NO: 27; or, comprises an amino acid sequence differing from the amino acid sequence shown at positions 16-228 or 17-216 of SEQ ID NO: 27 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with an identity of positions 16-228 or 17-216 of SEQ ID NO: 27, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0015] Preferably, the extracellular domain comprises 1-10 amino acid residues (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10) at the N-terminus and / or C-terminus of the extracellular domain of the non-human animal endogenous IL22RA1 protein, more preferably comprising the amino acid sequence shown at positions 16 and 217-230 of SEQ ID NO:26; or, comprising amino acid sequences with at least 77%, 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown at positions 16 and 217-230 of SEQ ID NO:26; or, comprising amino acid sequences with differences not exceeding 10, 9, 8, 7, 6, 5, 4, 3, 2, or not exceeding 1 amino acid from the amino acid sequence shown at positions 16 and 217-230 of SEQ ID NO:26; or, comprising amino acid sequences with differences not exceeding 10, 9, 8, 7, 6, 5, 4, 3, 2, or not exceeding 1 amino acid from the amino acid sequence shown at positions 16 and 217-230 of SEQ ID NO:26; or, comprising amino acid sequences with differences not exceeding 10, 9, 8, 7, 6, 5, 4, 3, 2, or not exceeding 1 amino acid from the amino acid sequence shown at positions 16 and 217-230 of SEQ ID NO:26. The amino acid sequence shown at positions 16 and 217-230 of NO:26 includes substitutions, deletions, and / or insertions of one or more amino acid residues.

[0016] Preferably, the non-human animal is a rodent, and more preferably a rat or mouse.

[0017] Preferably, the modified IL22RA1 protein contains a cytoplasmic domain that is substantially the same as that of the non-human endogenous IL22RA1 protein.

[0018] Preferably, the amino acid sequence of the cytoplasmic domain comprises the amino acid sequence shown in positions 252-581 of SEQ ID NO: 26; or, comprises an amino acid sequence with an identity of at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown in positions 252-581 of SEQ ID NO: 26; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 252-581 of SEQ ID NO: 26 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with an identity of positions 252-581 of SEQ ID NO: 26, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0019] Preferably, the modified IL22RA1 protein contains a transmembrane domain that is substantially the same as that of the non-human endogenous IL22RA1 protein.

[0020] Preferably, the amino acid sequence of the transmembrane domain comprises the amino acid sequence shown in positions 231-251 of SEQ ID NO: 26; or, comprises an amino acid sequence with an identity of at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown in positions 231-251 of SEQ ID NO: 26; or, comprises an amino acid sequence with a difference of no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid from the amino acid sequence shown in positions 231-251 of SEQ ID NO: 26; or, comprises an amino acid sequence with a substitution, deletion, and / or insertion of one or more amino acid residues as shown in positions 231-251 of SEQ ID NO: 26.

[0021] Preferably, the modified IL22RA1 protein has an amino acid sequence comprising the amino acid sequence shown in SEQ ID NO: 32; or, has an amino acid sequence identity with the amino acid sequence shown in SEQ ID NO: 32 of at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99%; or, has an amino acid sequence differing from the amino acid sequence shown in SEQ ID NO: 32 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, has an amino acid sequence similar to that shown in SEQ ID NO: 32, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0022] Preferably, the nucleotide sequence of the replaced segment encoding the corresponding region of non-human endogenous IL22RA1 includes all or part of the nucleotide sequence encoding the extracellular domain of the non-human endogenous IL22RA1 protein, more preferably including the nucleotide sequence encoding positions 17-216 or 16-230 of SEQ ID NO: 26, or including at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence encoding positions 17-216 or 16-230 of SEQ ID NO: 26; or including a difference of no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide from the nucleotide sequence encoding positions 17-216 or 16-230 of SEQ ID NO: 26; or including a difference of no more than 1 nucleotide from the nucleotide sequence encoding SEQ ID NO: 26. The nucleotide sequence shown in NO:26, from positions 17-216 or 16-230, includes a nucleotide sequence with substitutions, deletions, and / or insertions of one or more nucleotides. More preferably, it includes a partial nucleotide sequence of exons 2 to 5 of the non-human animal endogenous IL22RA1 sequence.

[0023] Preferably, the nucleotide sequence encoding the human IL22RA1 region comprises a portion of the human IL22RA1 gene; more preferably, it comprises all or part of exons 2 to 5 of the human IL22RA1 gene, and even more preferably, it comprises a portion of exon 2, all of exon 3, all of exon 4, and a portion of exon 5. Preferably, the nucleotide sequence encoding the human IL22RA1 region comprises portions of exons 2 to 5. The exon 2 portion contains at least 50 bp of nucleotide sequence, for example, at least 50, 70, 90, 100, 120, 125, 126, 127, 128, 129, 130, or 133 bp of nucleotide sequence; the exon 5 portion contains at least 50 bp of nucleotide sequence, for example, at least 50, 70, 100, 110, 115, 116, 117, 118, 119, 120, 130, or 139 bp of nucleotide sequence.

[0024] Preferably, the nucleotide sequence encoding the human IL22RA1 region comprises the nucleotide sequence shown in SEQ ID NO: 30; or, comprises a nucleotide sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 30; or, comprises a nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 30 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or, comprises a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 30, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0025] Preferably, the mRNA transcribed from the nucleotide sequence encoding the modified IL22RA1 protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 31; B) the nucleotide sequence having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 31; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 31 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 31, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0026] Preferably, the non-human animal genome includes replacing all or part of the nucleotide sequence encoding the extracellular domain of the endogenous IL22RA1 protein in the non-human animal genome with a nucleotide sequence encoding an extracellular domain substantially identical to that of the human IL22RA1 protein.

[0027] Preferably, the non-human animal genome includes replacing all or part of the nucleotide sequence encoding the extracellular domain of the endogenous IL22RA1 protein in the non-human animal genome with a nucleotide sequence encoding the human IL22RA1 region.

[0028] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding positions 17-216 or 16-230 of SEQ ID NO: 26 in the non-human animal genome with a nucleotide sequence encoding the human IL22RA1 region (e.g., genomic DNA sequence, cDNA sequence or CDS sequence).

[0029] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding positions 17-216 or 16-230 of SEQ ID NO: 26 in the non-human animal genome with portions of exons 2 to 5 of the human IL22RA1 gene.

[0030] Preferably, the non-human animal genome includes replacing parts of exons 2 to 5 of the non-human animal endogenous IL22RA1 gene in the non-human animal genome with parts of exons 2 to 5 of the human IL22RA1 gene.

[0031] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding positions 16-230 of SEQ ID NO: 26 in the non-human animal genome with the nucleotide sequence encoding positions 16-228 of SEQ ID NO: 27.

[0032] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding positions 17-216 of SEQ ID NO: 26 in the non-human animal genome with the nucleotide sequence encoding positions 17-216 of SEQ ID NO: 27.

[0033] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding SEQ ID NO: 26 in the non-human animal genome with the nucleotide sequence encoding SEQ ID NO: 32.

[0034] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding positions 17-216 of SEQ ID NO: 30 in the non-human animal genome with the nucleotide sequence shown in SEQ ID NO: 30.

[0035] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding SEQ ID NO: 26 in the non-human animal genome with the nucleotide sequence shown in SEQ ID NO: 31.

[0036] Preferably, the expression of the modified IL22RA1 protein is controlled by endogenous IL22RA1 regulatory elements (e.g., promoter, 5'UTR and / or 3'UTR).

[0037] Preferably, the non-human animal genome is homozygous or heterozygous for the genetically modified endogenous IL22RA1 gene.

[0038] Preferably, the non-human animal genome contains endogenous genes encoding human or modified or wild-type other genes, and preferably, the other genes are selected from at least one of IL22, IL10RB, IL20RA, IL20RB, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3 and OX40.

[0039] Preferably, the non-human animal genome contains an endogenous IL22 locus encoding a human, modified, or wild-type IL22 protein.

[0040] In one specific embodiment of the present invention, the non-human animal genome contains a genetically modified endogenous IL22 gene encoding a human or modified IL22 protein, and the non-human animal expresses the human or modified IL22 protein.

[0041] Preferably, in the genetically modified endogenous IL22 locus, the nucleotide sequence encoding the corresponding segment of non-human animal endogenous IL22 is replaced with the nucleotide sequence encoding the human IL22 segment.

[0042] Preferably, the human or modified IL22 protein comprises the amino acid sequence shown in SEQ ID NO: 2; or, comprises an amino acid sequence with at least 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in SEQ ID NO: 2; or, comprises an amino acid sequence differing from the amino acid sequence shown in SEQ ID NO: 2 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence similar to that shown in SEQ ID NO: 2, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0043] Preferably, the nucleotide sequence encoding the human IL22 region includes a portion of the human IL22 gene; more preferably, it includes a portion of exon 2, all of exon 3, all of exon 4, all of exon 5, and a portion of exon 6 of the human IL22 gene; even more preferably, it includes portions of exons 2 to 6.

[0044] Preferably, the nucleotide sequence encoding the human IL22 region comprises the nucleotide sequence shown in SEQ ID NO: 5; or, comprises a nucleotide sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 5; or, comprises a nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 5 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or, comprises a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 5, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0045] Preferably, the mRNA transcribed from the nucleotide sequence encoding the human or modified IL22 protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 6; B) the nucleotide sequence having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 6; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 6 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 6, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0046] Preferably, the non-human animal genome contains an endogenous IL10RB locus encoding a human, modified, or wild-type IL10RB protein.

[0047] In one specific embodiment of the present invention, the non-human animal genome contains a genetically modified endogenous IL10RB locus encoding a modified IL10RB protein, and the non-human animal expresses the modified IL10RB protein.

[0048] Preferably, in the genetically modified endogenous IL10RB locus, the nucleotide sequence encoding the corresponding segment of non-human endogenous IL10RB is replaced with the nucleotide sequence encoding the human IL10RB segment.

[0049] Preferably, the modified IL10RB protein comprises the amino acid sequence shown in SEQ ID NO: 62; or, comprises an amino acid sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in SEQ ID NO: 62; or, comprises an amino acid sequence differing from the amino acid sequence shown in SEQ ID NO: 62 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence similar to that shown in SEQ ID NO: 62, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0050] Preferably, the nucleotide sequence encoding the human IL10RB region includes a portion of the human IL10RB gene; more preferably, it includes a portion of exon 2, all of exon 3, all of exon 4, all of exon 5, and a portion of exon 6 of the human IL10RB gene; even more preferably, it includes portions of exons 2 to 6.

[0051] Preferably, the nucleotide sequence encoding the human IL10RB region comprises the nucleotide sequence shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or a variant thereof; or, comprises a nucleotide sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity to the nucleotide sequence shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or a variant thereof; or, comprises a nucleotide sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity to the nucleotide sequence shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or a variant thereof. The nucleotide sequence shown at positions 33268405-33288111 or its variants differ by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2 or no more than 1 nucleotide; or, a nucleotide sequence containing the nucleotide sequence shown at positions 33268405-33288111 or its variants with NCBI gene accession number NC_000021.9, including substitutions, deletions and / or insertions of one or more nucleotides, wherein the variants include one or more of the following: C>T mutation at position 33276503, G>A mutation at position 33276818, or TTTTGGG>TTG mutation at positions 33277048-33277053;

[0052] Preferably, the mRNA transcribed from the nucleotide sequence encoding the modified IL10RB protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 61; B) the nucleotide sequence having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 61; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 61 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 61, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0053] Preferably, the non-human animal genome contains an endogenous IL20RA locus encoding a human, modified, or wild-type IL20RA protein.

[0054] In one specific embodiment of the present invention, the non-human animal genome contains a genetically modified endogenous IL20RA locus encoding a modified IL20RA protein, and the non-human animal expresses the modified IL20RA protein.

[0055] Preferably, in the genetically modified endogenous IL20RA locus, the nucleotide sequence encoding the corresponding region of non-human endogenous IL20RA is replaced with the nucleotide sequence encoding the human IL20RA region.

[0056] Preferably, the modified IL20RA protein has an amino acid sequence comprising the amino acid sequence shown in SEQ ID NO: 91; or, having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in SEQ ID NO: 91; or, differing from the amino acid sequence shown in SEQ ID NO: 91 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, having an amino acid sequence similar to that shown in SEQ ID NO: 91, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0057] Preferably, the nucleotide sequence encoding the human IL20RA segment includes a portion of the human IL20RA gene; more preferably, it includes a portion of exon 2, all of exon 3, all of exon 4, all of exon 5, and a portion of exon 6 of the human IL20RA gene; even more preferably, it includes portions of exons 2 to 6.

[0058] Preferably, the nucleotide sequence encoding the human IL20RA region comprises the nucleotide sequence shown in positions 137004735-137017092 of NCBI Gene Accession Number NC_000006.12; or, it comprises a nucleotide sequence with an identity of at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99. 8% or at least 99% of the nucleotide sequence; or, a nucleotide sequence that differs from the nucleotide sequence shown in positions 137004735-137017092 of NCBI Gene Accession Number NC_000006.12 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2 or no more than 1 nucleotide; or, a nucleotide sequence that includes substitutions, deletions and / or insertions of one or more nucleotides as shown in the nucleotide sequence shown in positions 137004735-137017092 of NCBI Gene Accession Number NC_000006.12.

[0059] Preferably, the mRNA transcribed from the nucleotide sequence encoding the modified IL20RA protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 90; B) the nucleotide sequence having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 90; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 90 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 90, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0060] Preferably, the non-human animal genome contains an endogenous IL20RB locus encoding a human, modified, or wild-type IL20RB protein.

[0061] In one specific embodiment of the present invention, the non-human animal genome contains a genetically modified endogenous IL20RB locus encoding a modified IL20RB protein, and the non-human animal expresses the modified IL20RB protein.

[0062] Preferably, in the genetically modified endogenous IL20RB locus, the nucleotide sequence encoding the corresponding segment of non-human animal endogenous IL20RB is replaced with the nucleotide sequence encoding the human IL20RB segment.

[0063] Preferably, the modified IL20RB protein has an amino acid sequence comprising the amino acid sequence shown in SEQ ID NO: 121; or, having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in SEQ ID NO: 121; or, differing from the amino acid sequence shown in SEQ ID NO: 121 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, having an amino acid sequence similar to that shown in SEQ ID NO: 121, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0064] Preferably, the nucleotide sequence encoding the human IL20RB region includes a portion of the human IL20RB gene; more preferably, it includes a portion of exon 2, all of exon 3, all of exon 4, all of exon 5, and a portion of exon 6 of the human IL20RB gene; even more preferably, it includes portions of exons 2 to 6.

[0065] Preferably, the nucleotide sequence encoding the human IL20RB region comprises the nucleotide sequence shown in positions 136980507-136995463 of NCBI Gene Accession Number NC_000003.12; or, it comprises a nucleotide sequence with an identity of at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 9 ... 8% or at least 99% of the nucleotide sequence; or, a nucleotide sequence that differs from the nucleotide sequence shown in positions 136980507-136995463 of NCBI Gene Accession Number NC_000003.12 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2 or no more than 1 nucleotide; or, a nucleotide sequence that includes substitutions, deletions and / or insertions of one or more nucleotides as shown in the nucleotide sequence shown in positions 136980507-136995463 of NCBI Gene Accession Number NC_000003.12.

[0066] Preferably, the mRNA transcribed from the nucleotide sequence encoding the modified IL20RB protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 120; B) the nucleotide sequence having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 120; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 120 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 120, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0067] Preferably, the non-human animal genome contains an endogenous IL10 locus encoding a human, modified, or wild-type IL10 protein.

[0068] Preferably, the non-human animal or non-human animal genome contains a genetically modified endogenous IL10 locus encoding a human or modified IL10 protein, and the non-human animal expresses the human or modified IL10 protein.

[0069] Preferably, in the genetically modified endogenous IL10 locus, the nucleotide sequence of the corresponding segment of the non-human animal endogenous IL10 is replaced with the nucleotide sequence encoding the human IL10 segment.

[0070] Preferably, the nucleotide sequence encoding the human IL10 region comprises a portion of exon 1, all of exon 2, all of exon 3, all of exon 4, and a portion of exon 5 of the human IL10 gene; more preferably, it comprises portions of exons 1 to 5. The portion of exon 1 comprises at least 50 bp, for example, at least 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, 160, 165, 170, 180, 190, 200, 220, or 224 bp of continuous nucleotide sequence. The portion of exon 5 contains at least 50 bp, such as at least 50, 60, 70, 80, 90, 93, 100, 200, 300, 400, 500, 600, 700, 800, 900, 1000, 1100, 1120, or 1127 bp of continuous nucleotide sequence.

[0071] Preferably, the non-human animal genome includes replacing all or part of the nucleotide sequence encoding the endogenous IL10 protein in the non-human animal genome with the nucleotide sequence encoding all or part of the human IL10 protein.

[0072] Preferably, the non-human animal genome includes replacing all or part of the nucleotide sequence encoding the endogenous IL10 protein in the non-human animal genome with a nucleotide sequence encoding the human IL10 region.

[0073] Preferably, the non-human animal genome includes replacing parts of exons 1 to 5 of the non-human animal endogenous IL10 gene in the non-human animal genome with parts of exons 1 to 5 of the human IL10 gene.

[0074] Preferably, the expression of human or modified IL10 protein is controlled by endogenous IL10 regulatory elements (e.g., promoter, 5'UTR and / or 3'UTR).

[0075] Preferably, the non-human animal or non-human animal genome is homozygous or heterozygous for the genetically modified endogenous IL10 gene.

[0076] Preferably, the non-human animal genome contains an endogenous IL10RA locus encoding a human, modified, or wild-type IL10RA protein.

[0077] Preferably, the non-human animal or the non-human animal genome contains a genetically modified endogenous IL10RA locus encoding a modified IL10RA protein, and the non-human animal expresses the modified IL10RA protein.

[0078] Preferably, in the genetically modified endogenous IL10RA locus, the nucleotide sequence encoding the corresponding segment of non-human animal endogenous IL10RA is replaced with the nucleotide sequence encoding the human IL10RA segment.

[0079] Preferably, the modified IL10RA protein comprises a cytoplasmic domain, a transmembrane domain, and an extracellular domain.

[0080] Preferably, the modified IL10RA protein further comprises a signal peptide.

[0081] Preferably, the IL10RA protein contains a signal peptide that is substantially the same as that of the human IL10RA protein.

[0082] Preferably, the IL10RA protein contains an extracellular domain that is substantially the same as that of the human IL10RA protein.

[0083] Preferably, the IL10RA protein contains a transmembrane domain that is substantially the same as that of the human IL10RA protein.

[0084] Preferably, the IL10RA protein contains a cytoplasmic domain that is substantially the same as that of the human IL10RA protein.

[0085] Preferably, the cytoplasmic domain also includes all or part of the cytoplasmic domain of the non-human animal endogenous IL10RA protein.

[0086] Preferably, the nucleotide sequence encoding the human IL10RA segment includes a portion of exon 1, all of exon 2, all of exon 3, all of exon 4, all of exon 5, and a portion of exon 6 of the human IL10RA gene; more preferably, it includes portions of exons 1 to 6.

[0087] Preferably, the non-human animal genome contains a nucleotide sequence that replaces all or part of the extracellular domain encoding the endogenous IL10RA protein in the non-human animal genome with a nucleotide sequence encoding an extracellular domain substantially identical to that of the human IL10RA protein.

[0088] Preferably, the non-human animal genome contains nucleotide sequences that replace all or part of the signal peptide and extracellular domain encoding the endogenous IL10RA protein in the non-human animal genome with nucleotide sequences encoding signal peptides and extracellular domains that are substantially identical to those of the human IL10RA protein.

[0089] Preferably, the non-human animal genome includes nucleotide sequences that replace all or part of the signal peptide, extracellular domain, and transmembrane domain encoding the endogenous IL10RA protein in the non-human animal genome with nucleotide sequences encoding signal peptides, extracellular domains, and transmembrane domains that are substantially identical to those of the human IL10RA protein.

[0090] Preferably, the non-human animal genome includes replacing all or part of the nucleotide sequences encoding the endogenous IL10RA protein in the non-human animal genome with nucleotide sequences encoding signal peptides, extracellular domains, transmembrane domains, and cytoplasmic domains that are substantially identical to those of the human IL10RA protein.

[0091] Preferably, the non-human animal genome includes nucleotide sequences that replace all or part of the signal peptide, extracellular domain, transmembrane domain, and cytoplasmic domain encoding the endogenous IL10RA protein in the non-human animal genome with nucleotide sequences encoding the human IL10RA region.

[0092] Preferably, the expression of the modified IL10RA protein is controlled by endogenous IL10RA regulatory elements (e.g., promoters, 5'UTR, and / or 3'UTR).

[0093] Preferably, the non-human animal genome is homozygous or heterozygous for the genetically modified endogenous IL10RA gene.

[0094] Preferably, the non-human animal genome comprises any one of the following groups:

[0095] 1) Genetically modified endogenous IL22RA1 gene encoding modified IL22RA1 protein and wild-type endogenous IL22 gene;

[0096] 2) Genetically modified endogenous IL22RA1 genes encoding modified IL22RA1 protein, genetically modified endogenous IL22 genes encoding human or modified IL22 protein, and wild-type endogenous IL10RB genes;

[0097] 3) Genetically modified endogenous IL22RA1 gene and wild-type endogenous IL20RB gene encoding modified IL22RA1 protein;

[0098] 4) Genetically modified endogenous IL22RA1 gene, wild-type endogenous IL20RB gene, and wild-type endogenous IL20 gene encoding modified IL22RA1 protein;

[0099] 5) Genetically modified endogenous IL22RA1 gene encoding modified IL22RA1 protein, genetically modified endogenous IL20RB gene encoding modified IL20RB protein, and wild-type endogenous IL20 gene;

[0100] 6) Genetically modified endogenous IL22RA1 gene, wild-type endogenous IL20RB gene, and wild-type endogenous IL24 gene encoding modified IL22RA1 protein;

[0101] 7) Genetically modified endogenous IL22RA1 gene encoding modified IL22RA1 protein, genetically modified endogenous IL20RB gene encoding modified IL20RB protein, and wild-type endogenous IL24 gene.

[0102] 8) Genetically modified endogenous IL22RA1 gene encoding modified IL22RA1 protein, genetically modified endogenous IL10RB gene encoding modified IL10RB protein, and wild-type endogenous IL22 gene.

[0103] Preferably, in non-human animals containing the non-human animal genome, the differentiation, development, and distribution of leukocyte subtypes and / or T cell subtypes are not significantly different from those in non-human animals expressing wild-type endogenous IL22, wild-type endogenous IL22RA1, and wild-type endogenous IL10RB.

[0104] Among them, white blood cell subtypes include one or more of B cells, T cells, NK cells, dendritic cells (DCs), granulocytes, macrophages, or monocytes. T cell subtypes include one or more of CD4+ T cells, CD8+ T cells, or Tregs.

[0105] Preferably, the leukocytes or T cells include leukocytes or T cells derived from the spleen, blood, or lymph nodes.

[0106] Preferably, in non-human animals containing the non-human animal genome, the expression levels of CXCL1 and IL6 are not significantly different from those in non-human animals expressing wild-type endogenous IL22, wild-type endogenous IL10RB, and wild-type endogenous IL22RA1.

[0107] In a second aspect, the present invention provides a non-human animal comprising a genetically modified endogenous IL22RA1 gene encoding a modified IL22RA1 protein, wherein the non-human animal expresses the modified IL22RA1 protein.

[0108] Preferably, the relevant limitations on the modified IL22RA1 protein and the genetically modified endogenous IL22RA1 gene are the same as those in the first aspect of the present invention.

[0109] Preferably, the non-human animal comprises the non-human animal genome described in the first aspect.

[0110] A third aspect of the present invention provides a method for preparing a non-human animal, wherein the non-human animal or the genome of the non-human animal contains a genetically modified endogenous IL22RA1 gene encoding a modified IL22RA1 protein, and the non-human animal expresses the modified IL22RA1 protein.

[0111] Preferably, the genome of the non-human animal or non-human animal contains a nucleotide sequence that replaces the nucleotide sequence encoding the corresponding segment of the non-human animal endogenous IL22RA1 with a nucleotide sequence encoding the human IL22RA1 segment.

[0112] Preferably, the modified IL22RA1 protein comprises a cytoplasmic domain, a transmembrane domain, and an extracellular domain.

[0113] Preferably, the modified IL22RA1 protein contains an extracellular domain that is substantially the same as that of the human IL22RA1 protein;

[0114] Preferably, all or part of the extracellular domain comprises the amino acid sequence shown at positions 16-228 or 17-216 of SEQ ID NO: 27; or, comprises an amino acid sequence with an identity of at least 77%, 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown at positions 16-228 or 17-216 of SEQ ID NO: 27; or, comprises an amino acid sequence differing from the amino acid sequence shown at positions 16-228 or 17-216 of SEQ ID NO: 27 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with an identity of positions 16-228 or 17-216 of SEQ ID NO: 27, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0115] Preferably, the non-human animal is a rodent, and more preferably a rat or mouse.

[0116] Preferably, the modified IL22RA1 protein contains a cytoplasmic domain that is substantially the same as that of the non-human endogenous IL22RA1 protein.

[0117] Preferably, the amino acid sequence of the cytoplasmic domain comprises the amino acid sequence shown in positions 252-581 of SEQ ID NO: 26; or, comprises an amino acid sequence with at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in positions 252-581 of SEQ ID NO: 26; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 252-581 of SEQ ID NO: 26 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with substitutions, deletions, and / or insertions of one or more amino acid residues as shown in positions 252-581 of SEQ ID NO: 26.

[0118] Preferably, the modified IL22RA1 protein contains a transmembrane domain that is substantially the same as that of the non-human endogenous IL22RA1 protein.

[0119] Preferably, the amino acid sequence of the transmembrane domain comprises the amino acid sequence shown in positions 231-251 of SEQ ID NO: 26; or, comprises an amino acid sequence with an identity of at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown in positions 231-251 of SEQ ID NO: 26; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 231-251 of SEQ ID NO: 26 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with an identity of positions 231-251 of SEQ ID NO: 26, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0120] Preferably, the modified IL22RA1 protein has an amino acid sequence comprising the amino acid sequence shown in SEQ ID NO: 32; or, having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in SEQ ID NO: 32; or, differing from the amino acid sequence shown in SEQ ID NO: 32 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, having an amino acid sequence similar to that shown in SEQ ID NO: 32, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0121] Preferably, the nucleotide sequence encoding the human IL22RA1 region includes a portion of exon 2, all of exon 3, all of exon 4, and a portion of exon 5 of the human IL22RA1 gene.

[0122] Preferably, the nucleotide sequence encoding the human IL22RA1 region comprises the nucleotide sequence shown in SEQ ID NO: 30; or, comprises a nucleotide sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 30; or, comprises a nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 30 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or, comprises a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 30, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0123] Preferably, the mRNA transcribed from the nucleotide sequence encoding the modified IL22RA1 protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 31; B) the nucleotide sequence having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 31; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 31 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 31, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0124] Preferably, the expression of the modified IL22RA1 protein is controlled by endogenous IL22RA1 regulatory elements (e.g., promoter, 5'UTR and / or 3'UTR).

[0125] Preferably, the genome of the non-human animal is homozygous or heterozygous for the genetically modified endogenous IL22RA1 gene.

[0126] Preferably, the genome of the non-human animal or the non-human animal contains an endogenous gene encoding a human or modified other gene. Preferably, the other gene is selected from at least one of IL22, IL10RB, IL20RA, IL20RB, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3, and OX40.

[0127] In a fourth aspect, the present invention provides a nucleic acid comprising a nucleotide sequence encoding an IL22RA1 protein modified as described above. Preferably, the nucleic acid further comprises SEQ ID NO: 30. Preferably, the mRNA transcribed from the nucleic acid comprises any of the following: A) the nucleotide sequence shown in SEQ ID NO: 31; B) having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 31; C) differing from the nucleotide sequence shown in SEQ ID NO: 31 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) having the nucleotide sequence shown in SEQ ID NO: 31, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0128] In a fifth aspect, the present invention provides a polypeptide encoded by the genetically modified endogenous IL22RA1 gene or the aforementioned nucleic acid.

[0129] In a sixth aspect, the present invention provides a humanized IL22RA1 protein, said humanized IL22RA1 protein comprising all or part of the human IL22RA1 protein.

[0130] Preferably, the humanized IL22RA1 protein comprises all or part of the signal peptide, extracellular domain, transmembrane domain and / or cytoplasmic domain of the human IL22RA1 protein.

[0131] In one specific embodiment of the present invention, the humanized IL22RA1 protein comprises all or part of the extracellular domain of the human IL22RA1 protein, preferably comprising at least 50 consecutive amino acids of the extracellular domain of the human IL22RA1 protein, for example comprising at least 50, 70, 90, 100, 150, 170, 190, 200, 201, 202, 203, 204, 205, 210, and 213 consecutive amino acids.

[0132] Preferably, the humanized IL22RA1 protein portion comprises the amino acid sequence shown in positions 17-216 of SEQ ID NO: 27; or, comprises an amino acid sequence with an identity of at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown in positions 17-216 of SEQ ID NO: 27; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 17-216 of SEQ ID NO: 27 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with an identity of positions 17-216 of SEQ ID NO: 27, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0133] Preferably, the humanized IL22RA1 protein comprises all or part of the amino acid sequence encoding exons 1 to 7 of the human IL22RA1 gene. More preferably, it comprises all or part of the amino acid sequence encoding any one, two, three or more, or a combination of two or more consecutive exons 1 to 7. More preferably, it comprises all or part of the amino acid sequence encoding exons 2 to 5. Even more preferably, it comprises a portion of exon 2, all of exons 3 and 4, and a portion of exon 5. In one specific embodiment of the invention, the humanized IL22RA1 protein comprises a portion of the amino acid sequence encoding exons 2 to 5 of the human IL22RA1 gene. The exon 2 portion contains at least a 50 bp nucleotide sequence, for example, at least 50, 70, 90, 100, 120, 125, 126, 127, 128, 129, 130, or 133 bp nucleotide sequences; preferably, the exon 2 portion contains a nucleotide sequence from the first to fifth (e.g., 1, 2, 3, 4, 5) amino acids encoding the N-terminus of the extracellular domain to the last nucleotide of the exon 2 portion, containing a nucleotide sequence encoding at least five consecutive amino acids of the extracellular domain of the human IL22RA1 protein, for example, a nucleotide sequence containing at least 5, 7, 9, 10, 15, 20, 25, 30, 35, 40, 43, or 44 consecutive amino acids. Preferably, the exon 5 portion contains at least a 50 bp nucleotide sequence, for example, at least 50, 70, 100, 110, 115, 116, 117, 118, 119, 120, 130, or 139 bp nucleotide sequences; preferably, the exon 5 portion contains a nucleotide sequence from the first nucleotide of exon 5 to 6-15 amino acids (e.g., 6, 7, 8, 9, 10, 11, 12, 13, 14, 15) encoding the C-terminus of the extracellular domain, and contains a nucleotide sequence encoding at least 5 consecutive amino acids of the extracellular domain of human IL22RA1 protein, for example, a nucleotide sequence containing at least 5, 7, 9, 10, 15, 20, 25, 30, 35, 39, 40, 45, or 46 consecutive amino acids.

[0134] Preferably, the humanized IL22RA1 protein further comprises all or part of the non-human animal IL22RA1 protein, and more preferably all or part of the signal peptide, extracellular domain, transmembrane domain and / or cytoplasmic domain of the non-human animal IL22RA1 protein.

[0135] In one specific embodiment of the present invention, the humanized IL22RA1 protein further comprises all or part of the non-human animal IL22RA1 protein, preferably comprising the amino acid sequence shown in positions 1-16 and 217-581 of SEQ ID NO: 26; or, comprising an amino acid sequence with an identity of at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with ...10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, or at least 1 amino acid with the amino acid sequence shown in positions 1-16 and 217-581 of SEQ ID NO: 26; or, comprising an amino acid sequence with an identity of at least 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, or at least 1 amino acid with the amino acid sequence shown in positions 1-16 and 217-581 of SEQ ID NO: 26; or, comprising an amino acid sequence with an identity of at least 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, or at least 1 amino acid with the amino acid sequence shown in positions The amino acid sequence shown in NO:26, positions 1-16 and 217-581, includes substitutions, deletions, and / or insertions of one or more amino acid residues.

[0136] In one specific embodiment of the present invention, the amino acid sequence of the humanized IL22RA1 protein comprises any one of the following groups: A) the amino acid sequence shown in SEQ ID NO: 32; B) the amino acid sequence being at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identical to the amino acid sequence shown in SEQ ID NO: 32; C) the amino acid sequence differing from the amino acid sequence shown in SEQ ID NO: 32 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or D) the amino acid sequence shown in SEQ ID NO: 32, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0137] Preferably, the non-human animal can be selected from rodents, pigs, rabbits, monkeys, or any other non-human animal that can be genetically edited to produce humanized genes. Preferably, the non-human animal is a non-human mammal. More preferably, the non-human mammal is a rodent. Even more preferably, the rodent is a rat or mouse. Preferably, the non-human animal is an immunodeficient non-human mammal. More preferably, the immunodeficient non-human mammal is an immunodeficient rodent, an immunodeficient pig, an immunodeficient rabbit, or an immunodeficient monkey. Even more preferably, the immunodeficient rodent is an immunodeficient mouse or rat. Most preferably, the immunodeficient mouse is a NOD-Prkdc mouse. scid IL-2rγ null Mice, NOD-Rag 1 - / - -IL2rg - / - (NRG) mice, Rag 2- / - -IL2rg - / - (RG) mice, NOD / SCID mice, or nude mice.

[0138] In a seventh aspect, the present invention provides a humanized IL22RA1 gene, wherein the humanized IL22RA1 gene comprises a portion of the human IL22RA1 gene.

[0139] Preferably, the humanized IL22RA1 gene comprises all or part of exons 1 to 7 of the human IL22RA1 gene. More preferably, the humanized IL22RA1 gene comprises all or part of any one, two, three or more, or consecutive combinations of two or more exons 1 to 7 of the human IL22RA1 gene. More preferably, it comprises all or part of exons 2 to 5. Even more preferably, the humanized IL22RA1 gene comprises a portion of exon 2, all of exons 3 and 4, and a portion of exon 5 of the human IL22RA1 gene, preferably also comprising intron 2 and / or intron 4, and more preferably comprising any intron between exons 2 and 5. In one specific embodiment of the present invention, the humanized IL22RA1 gene comprises a portion of exon 2 to exon 5 of the human IL22RA1 gene, wherein the portion of exon 2 comprises at least 50 bp of nucleotide sequence, for example, at least 50, 70, 90, 100, 120, 125, 126, 127, 128, 129, 130, or 133 bp of nucleotide sequence; preferably, the portion of exon 2 comprises a nucleotide sequence from the first to fifth (e.g., 1, 2, 3, 4, 5) amino acids encoding the N-terminus of the extracellular domain to the last nucleotide of exon 2, comprising a nucleotide sequence encoding at least five consecutive amino acids of the extracellular domain of the human IL22RA1 protein, for example, a nucleotide sequence comprising at least 5, 7, 9, 10, 15, 20, 25, 30, 35, 40, 43, or 44 consecutive amino acids. Preferably, the exon 5 portion contains at least a 50 bp nucleotide sequence, for example, at least 50, 70, 100, 110, 115, 116, 117, 118, 119, 120, 130, or 139 bp nucleotide sequences; preferably, the exon 5 portion contains a nucleotide sequence from the first nucleotide of exon 5 to 6-15 amino acids (e.g., 6, 7, 8, 9, 10, 11, 12, 13, 14, 15) encoding the C-terminus of the extracellular domain, and contains a nucleotide sequence encoding at least 5 consecutive amino acids of the extracellular domain of human IL22RA1 protein, for example, a nucleotide sequence containing at least 5, 7, 9, 10, 15, 20, 25, 30, 35, 39, 40, 45, or 46 consecutive amino acids.

[0140] In one specific embodiment of the present invention, the humanized IL22RA1 gene comprises the nucleotide sequence shown in SEQ ID NO: 30; or, comprises a nucleotide sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 30; or, comprises a nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 30 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or, comprises a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 30, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0141] Preferably, the humanized IL22RA1 gene contains all or part of the nucleotide sequence encoding the human IL22RA1 protein. Further preferred are nucleotide sequences comprising all or part of the extracellular domain encoding the human IL22RA1 protein, wherein the humanized IL22RA1 gene comprises the nucleotide sequence encoding positions 17-216 of SEQ ID NO: 27; or, comprises a nucleotide sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity to the nucleotide sequence encoding positions 17-216 of SEQ ID NO: 27; or, comprises a nucleotide sequence differing from the amino acid sequence encoding positions 17-216 of SEQ ID NO: 27 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises a nucleotide sequence having the nucleotide sequence encoding positions 17-216 of SEQ ID NO: 27, including nucleotide sequences with substitutions, deletions, and / or insertions of one or more nucleotides.

[0142] Preferably, the humanized IL22RA1 gene further comprises all or part of the non-human animal IL22RA1 gene. More preferably, it comprises all or part of exons 1 to 7 of the non-human animal IL22RA1 gene, and more preferably, it comprises all of exon 1, part of exon 2, part of exon 5, and / or all of exons 6 to 7 of the non-human animal IL22RA1 gene.

[0143] In one specific embodiment of the present invention, the humanized IL22RA1 gene further comprises a nucleotide sequence encoding positions 1-16 and 217-581 of SEQ ID NO: 26; or, comprises a nucleotide sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity to the nucleotide sequence encoding positions 1-16 and 217-581 of SEQ ID NO: 26; or, comprises a nucleotide sequence differing from the amino acid sequence encoding positions 1-16 and 217-581 of SEQ ID NO: 26 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises a nucleotide sequence having a nucleotide sequence encoding positions 1-16 and 217-581 of SEQ ID NO: 26, including nucleotide sequences with substitutions, deletions, and / or insertions of one or more nucleotides.

[0144] In one specific embodiment of the present invention, the nucleotide sequence of the humanized IL22RA1 gene comprises any one of the following groups: A) the transcribed mRNA is the nucleotide sequence shown in SEQ ID NO: 31; B) the transcribed mRNA is at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identical to the nucleotide sequence shown in SEQ ID NO: 31; C) the transcribed mRNA differs from the nucleotide sequence shown in SEQ ID NO: 31 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) the transcribed mRNA has the nucleotide sequence shown in SEQ ID NO: 31, including nucleotide sequences with substitutions, deletions, and / or insertions of one or more nucleotides.

[0145] Preferably, the humanized IL22RA1 gene encodes the aforementioned humanized IL22RA1 protein.

[0146] Preferably, the humanized IL22RA1 gene further includes a specific inducer or repressor. More preferably, the specific inducer or repressor can be a conventionally induced or repressible substance. In one specific embodiment of the present invention, the specific inducer is selected from the tetracycline system (Tet-Off System / Tet-On System) or the tamoxifen system.

[0147] Preferably, the non-human animal can be selected from rodents, pigs, rabbits, monkeys, or any other non-human animal that can be genetically edited to produce humanized genes. Further limitations on the type of non-human animal are the same as in the sixth aspect of this invention.

[0148] In an eighth aspect, the present invention provides a targeting vector comprising a donor nucleotide sequence. Preferably, the donor nucleotide sequence comprises one of the following: A) a nucleotide sequence encoding a humanized IL22RA1 protein; B) a nucleotide sequence encoding all or part of a human IL22RA1 protein, preferably comprising all or part of a nucleotide sequence encoding a signal peptide, extracellular domain, transmembrane domain, and / or cytoplasmic domain of a human IL22RA1 protein, more preferably comprising all or part of a nucleotide sequence encoding an extracellular domain of a human IL22RA1 protein, more preferably comprising a nucleotide sequence encoding at least 50 consecutive amino acids of an extracellular domain of a human IL22RA1 protein, and even more preferably comprising an amino acid sequence encoding positions 17-216 of SEQ ID NO: 27; C) a nucleotide sequence of a humanized IL22RA1 gene; or D) a portion of a human IL22RA1 gene, preferably comprising all or part of exons 1 to 7 of the human IL22RA1 gene. Preferably, it contains all or part of any one, two, three or more, or consecutive combinations of two or more exons from exons 1 to 7. More preferably, it contains all or part of exons 2 to 5 of the human IL22RA1 gene; even more preferably, it contains a portion of exon 2, all of exons 3 and 4, and a portion of exon 5; more preferably, it contains a portion of exons 2 to 5, wherein the portion of exon 2 contains at least 50 bp of nucleotide sequence, for example, at least 50, 70, 90, 100, 120, 125, 126, 127, 128, 129, 130, etc. A 133 bp nucleotide sequence; preferably, the exon 2 portion comprises a nucleotide sequence from the first to the fifth (e.g., 1, 2, 3, 4, 5) amino acids encoding the N-terminus of the extracellular domain to the last nucleotide of exon 2, and a nucleotide sequence encoding at least five consecutive amino acids encoding the extracellular domain of human IL22RA1 protein, for example, a nucleotide sequence containing at least 5, 7, 9, 10, 15, 20, 25, 30, 35, 40, 43, 44 consecutive amino acids. Preferably, the exon 5 portion contains at least a 50 bp nucleotide sequence, for example, at least 50, 70, 100, 110, 115, 116, 117, 118, 119, 120, 130, or 139 bp nucleotide sequences; preferably, the exon 5 portion contains a nucleotide sequence from the first nucleotide of exon 5 to 6-15 amino acids (e.g., 6, 7, 8, 9, 10, 11, 12, 13, 14, 15) encoding the C-terminus of the extracellular domain, and contains a nucleotide sequence encoding at least 5 consecutive amino acids of the extracellular domain of human IL22RA1 protein, for example, a nucleotide sequence containing at least 5, 7, 9, 10, 15, 20, 25, 30, 35, 39, 40, 45, or 46 consecutive amino acids.Further preferably, the donor nucleotide sequence comprises the nucleotide sequence shown in SEQ ID NO: 30; or, comprises a nucleotide sequence with an identity of at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the nucleotide sequence shown in SEQ ID NO: 30; or, comprises a nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 30 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or, comprises a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 30, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0149] Preferably, the humanized IL22RA1 protein is the aforementioned humanized IL22RA1 protein.

[0150] Preferably, the humanized IL22RA1 gene is the aforementioned humanized IL22RA1 gene.

[0151] Preferably, the targeting vector further comprises a 5' arm (5' homologous arm) and / or a 3' arm (3' homologous arm).

[0152] The 5' arm is a DNA fragment homologous to the 5' end of the conversion region to be altered, selected from 100-10000 nucleotides in length from the genomic DNA of the non-human animal IL22RA1 gene. Preferably, the 5' arm is a nucleotide with at least 90% homology to NCBI accession number NC_000070.7. More preferably, the 5' arm sequence comprises SEQ ID NO: 28.

[0153] The 3' arm is a DNA fragment homologous to the 3' end of the conversion region to be altered, and is selected from 100-10000 nucleotides in length from the genomic DNA of the non-human animal IL22RA1 gene; preferably, the 3' arm is a nucleotide with at least 90% homology to NCBI accession number NC_000070.7; more preferably, the 3' arm sequence contains SEQ ID NO: 29.

[0154] Preferably, the targeting vector further comprises SEQ ID NO: 33 and / or 34.

[0155] Preferably, the conversion region to be altered is located on exons 2 to 5 of the non-human animal IL22RA1 gene.

[0156] Preferably, the targeting vector further comprises a marker gene. More preferably, the marker gene is a gene encoding a negative selection marker. Even more preferably, the gene encoding the negative selection marker is a gene encoding the diphtheria toxin A subunit (DTA). In a specific embodiment of the present invention, the targeting vector further comprises an resistance gene for positive clone selection. More preferably, the resistance gene for positive clone selection is the neomycin phosphotransferase coding sequence Neo. In a specific embodiment of the present invention, the targeting vector further comprises a specific recombination system. More preferably, the specific recombination system is a Frt recombination site (or a conventional loxP recombination system). There are two specific recombination systems, respectively mounted on both sides of the resistance gene.

[0157] A ninth aspect of the present invention provides a cell comprising the above-described targeting vector.

[0158] In a tenth aspect, the present invention provides the application of the above-described targeting vector and / or the above-described cells in IL22RA1 gene editing, preferably, the application includes, but is not limited to, knockout, insertion or replacement.

[0159] In an eleventh aspect, the present invention provides a non-human animal with humanized IL22RA1 gene, wherein the non-human animal expresses human or humanized IL22RA1 protein.

[0160] Preferably, the humanized IL22RA1 protein described above is expressed in the non-human animal.

[0161] Preferably, the expression of endogenous IL22RA1 protein in the non-human animal is reduced or absent.

[0162] Preferably, the non-human animal genome contains a portion of the human IL22RA1 gene or a humanized IL22RA1 gene, more preferably it contains the aforementioned humanized IL22RA1 gene.

[0163] Preferably, a portion of the human IL22RA1 gene or the nucleotide sequence of the humanized IL22RA1 gene is operatively linked to an endogenous regulatory element at an endogenous IL22RA1 gene locus in at least one chromosome.

[0164] Preferably, the non-human animal is constructed by introducing a donor nucleotide sequence into the non-human animal IL22RA1 locus. Preferably, the limitation of the donor nucleotide sequence is the same as that in aspect eight of the present invention.

[0165] Preferably, the non-human animal is constructed using the aforementioned targeting vector.

[0166] According to some embodiments of the present invention, the non-human animal further comprises other gene modifications, said other genes being selected from at least one of IL22, IL10RB, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3, and OX40.

[0167] Preferably, the IL22 gene is a humanized IL22 gene; more preferably, the humanized IL22 gene is any of the humanized IL22 genes described in this invention.

[0168] Preferably, the IL10RB gene is a humanized IL10RB gene; more preferably, the humanized IL10RB gene is any of the humanized IL10RB genes described in this invention.

[0169] Preferably, the IL10RB gene is a humanized IL20RA gene, and more preferably, the humanized IL20RA gene is any of the humanized IL20RA genes described in this invention.

[0170] Preferably, the IL10RB gene is a humanized IL20RB gene, and more preferably, the humanized IL20RB gene is any of the humanized IL20RB genes described in this invention.

[0171] According to some embodiments of the present invention, the partial or humanized IL22RA1 gene of the human IL22RA1 gene and / or the other genes are homozygous or heterozygous for the endogenously replaced locus.

[0172] Preferably, the humanized IL22RA1 gene further includes a specific inducer or repressor; more preferably, the specific inducer or repressor can be a conventional substance that can induce or repress.

[0173] In one specific embodiment of the present invention, the specific inducer is selected from the tetracycline system (Tet-Off System / Tet-On System) or the tamoxifen system.

[0174] Preferably, the non-human animal can be selected from rodents, pigs, rabbits, monkeys, or any other non-human animal that can be genetically edited to produce humanized genes. Further limitations on the type of non-human animal are the same as in the sixth aspect of this invention.

[0175] In a twelfth aspect, the present invention provides a method for constructing a non-human animal with humanized IL22RA1 gene, wherein the non-human animal expresses human or humanized IL22RA1 protein.

[0176] Preferably, the humanized IL22RA1 protein described above is expressed in the non-human animal.

[0177] Preferably, the genome of at least one cell of the non-human animal contains a portion of the human IL22RA1 gene or a humanized IL22RA1 gene, more preferably the aforementioned humanized IL22RA1 gene.

[0178] Preferably, the non-human animal is a non-human animal with the IL22RA1 gene humanized as described above.

[0179] Preferably, the construction method includes introducing a donor nucleotide sequence into the IL22RA1 gene locus in a non-human animal. Preferably, the limitation of the donor nucleotide sequence is the same as in aspect eight of the present invention.

[0180] Preferably, the human IL22RA1 gene, either partially or humanized, is operatively linked to an endogenous regulatory element at at least one endogenous IL22RA1 gene locus on a chromosome.

[0181] Preferably, the partial or humanized IL22RA1 gene of the human IL22RA1 gene is regulated in non-human animals by a regulatory element. More preferably, the regulatory element can be endogenous or exogenous.

[0182] Preferably, the control elements include, but are not limited to, a promoter, a 5'UTR, and / or a 3'UTR.

[0183] In one specific embodiment of the present invention, the endogenous regulatory element is derived from the non-human animal IL22RA1 gene, and the exogenous regulatory element is derived from the human IL22RA1 gene.

[0184] Preferably, the insertion site is located after the endogenous regulatory element of the IL22RA1 gene.

[0185] Preferably, the introduction is a substitution, insertion, or transgenesis. Optionally, the introduction of the non-human animal IL22RA1 gene locus is a substitution of the corresponding region in the non-human animal. More preferably, all or part of exons 2 to 5 of the non-human animal IL22RA1 gene is substituted. More preferably, part of exon 2, all of exons 3 to 4, and part of exon 5 of the non-human animal IL22RA1 gene are substituted. Even more preferably, part of exons 2 to 5 of the non-human animal IL22RA1 gene are substituted.

[0186] Preferably, the nucleotide sequence encoding the amino acids shown in positions 17-216 of SEQ ID NO:26 is replaced.

[0187] Preferably, the non-human animal used to humanize the IL22RA1 gene is constructed using gene editing technology, wherein the gene editing technology includes gene targeting technology using embryonic stem cells, CRISPR / Cas9 technology, zinc finger nuclease technology, transcription activator-like effector nuclease technology, homing endonuclease or other molecular biology techniques.

[0188] In one specific embodiment of the present invention, the construction method includes modifying the coding frame of the non-human animal IL22RA1 gene by inserting a nucleotide sequence encoding the human or humanized IL22RA1 protein, or a portion of the human IL22RA1 gene or a portion of the humanized IL22RA1 gene, after the endogenous regulatory element of the non-human animal IL22RA1 gene. The modified coding frame of the non-human animal IL22RA1 gene can be achieved by knocking out the functional region of the non-human animal IL22RA1 gene or by inserting a sequence that causes the non-human animal IL22RA1 protein to be not expressed, expressed at a reduced level, or expressed as a non-functional protein. More preferably, the modified coding frame of the non-human animal IL22RA1 gene can be a knockout of all or part of exons 2 to 5 of the non-human animal IL22RA1 gene, preferably a portion of exon 2, all of exon 3, all of exon 4, and a portion of exon 5, and even more preferably a portion of exons 2 to 5.

[0189] Preferably, the above-mentioned targeting vector is used to construct non-human animals.

[0190] In one specific embodiment of the present invention, the construction method includes introducing the above-mentioned targeting vector into non-human animal cells (preferably embryonic stem cells), screening out the correct positive clone cells and introducing them into the isolated blastocyst, culturing the blastocyst, and then transplanting the cultured blastocyst into the oviduct of a female non-human animal, allowing it to develop, and identifying and screening to obtain non-human animals with humanized IL22RA1 gene.

[0191] According to some embodiments of the present invention, the construction method further includes: mating non-human animals with humanized IL22RA1 genes with other genetically modified non-human animals, in vitro fertilization or direct gene editing, and screening to obtain multi-gene-modified non-human animals.

[0192] Preferably, the other genes are non-human animals modified with at least one of the following genes: IL22, IL10RB, IL20RA, IL20RB, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3, and OX40.

[0193] Preferably, the non-human animal also expresses at least one of the following human or humanized proteins: IL22, IL10RB, IL20RA, IL20RB, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3, and OX40.

[0194] Preferably, the IL22 gene is a humanized IL22 gene; more preferably, the humanized IL22 gene is any of the humanized IL22 genes described in this invention.

[0195] Preferably, the IL10RB gene is a humanized IL10RB gene; more preferably, the humanized IL10RB gene is any of the humanized IL10RB genes described in this invention.

[0196] Preferably, the IL10RB gene is a humanized IL20RA gene, and more preferably, the humanized IL20RA gene is any of the humanized IL20RA genes described in this invention.

[0197] Preferably, the IL10RB gene is a humanized IL20RB gene, and more preferably, the humanized IL20RB gene is any of the humanized IL20RB genes described in this invention.

[0198] Preferably, the IL22 protein is a humanized IL22 protein; more preferably, the humanized IL22 protein is any of the humanized IL22 proteins described in this invention.

[0199] Preferably, the IL10RB protein is a humanized IL10RB protein, and more preferably, the humanized IL10RB protein is any of the humanized IL10RB proteins described in this invention.

[0200] Preferably, the IL10RB protein is a humanized IL20RA protein, and more preferably, the humanized IL20RA protein is any of the humanized IL20RA proteins described in this invention.

[0201] Preferably, the IL10RB protein is a humanized IL20RB protein, and more preferably, the humanized IL20RB protein is any of the humanized IL20RB proteins described in this invention.

[0202] Preferably, in the genome of the multi-gene modified non-human animal, each of the modified genes is homozygous or heterozygous for the endogenous replaced locus.

[0203] Preferably, the non-human animal can be selected from rodents, pigs, rabbits, monkeys, or any other non-human animal that can be genetically edited to produce humanized genes. Further limitations on the type of non-human animal are the same as in the sixth aspect of this invention.

[0204] In a thirteenth aspect of the present invention, a non-human animal genome is provided, wherein the non-human animal or the non-human animal genome comprises a genetically modified endogenous IL22 gene encoding a human or modified IL22 protein, and the non-human animal expresses the human or modified IL22 protein.

[0205] Preferably, the non-human animal or non-human animal genome contains a nucleotide sequence encoding the corresponding segment of non-human animal endogenous IL22, which is replaced by a nucleotide sequence encoding the human IL22 segment.

[0206] Preferably, the human or modified IL22 protein comprises the amino acid sequence shown in SEQ ID NO: 2; or, comprises an amino acid sequence with at least 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in SEQ ID NO: 2; or, comprises an amino acid sequence differing from the amino acid sequence shown in SEQ ID NO: 2 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence similar to that shown in SEQ ID NO: 2, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0207] Preferably, the nucleotide sequence encoding the human IL22 region comprises a portion of the human IL22 gene; more preferably, it comprises a portion of exon 2, all of exon 3, all of exon 4, all of exon 5, and a portion of exon 6 of the human IL22 gene; more preferably, it comprises a portion of exons 2 to 6 of the human IL22 gene, wherein the exon 2 portion comprises at least 50 bp of continuous nucleotide sequence, for example, at least 50, 70, 90, 100, 150, 170, 180, 185, 186, 1... The nucleotide sequences are 87, 188, 189, 190, 200, and 232 bp. Preferably, the exon 2 portion contains from the start codon to the last nucleotide of exon 2, and the exon 6 portion contains at least 20 bp of continuous nucleotide sequences, such as at least 20, 30, 40, 50, 70, 75, 76, 77, 78, 79, 80, 100, 200, 500, 600, and 635 bp. Preferably, the exon 6 portion contains from the first nucleotide of exon 6 to the stop codon.

[0208] Preferably, the nucleotide sequence encoding the human IL22 region comprises the nucleotide sequence shown in SEQ ID NO: 5; or, comprises a nucleotide sequence with an identity of at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the nucleotide sequence shown in SEQ ID NO: 5; or, comprises a nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 5 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or, comprises a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 5, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0209] Preferably, the mRNA transcribed from the nucleotide sequence encoding the human or modified IL22 protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 6; B) the nucleotide sequence having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 6; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 6 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 6, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0210] Preferably, the nucleotide sequence of the replaced segment encoding the corresponding region of non-human endogenous IL22 includes a nucleotide sequence encoding the non-human endogenous IL22 protein, more preferably including the nucleotide sequence encoding SEQ ID NO: 1, or including at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence encoding SEQ ID NO: 1; or, including a nucleotide sequence differing from the nucleotide sequence encoding SEQ ID NO: 1 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or, including a nucleotide sequence having the nucleotide sequence encoding SEQ ID NO: 1, including substitution, deletion, and / or insertion of one or more nucleotides, more preferably including portions of exons 1 to 5 of the non-human endogenous IL22 sequence.

[0211] Preferably, the non-human animal genome includes replacing all or part of the nucleotide sequence encoding the endogenous IL22 protein in the non-human animal genome with a nucleotide sequence encoding all or part of human IL22.

[0212] Preferably, the non-human animal genome includes replacing all or part of the nucleotide sequence encoding the endogenous IL22 protein in the non-human animal genome with a nucleotide sequence encoding the human IL22 region.

[0213] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding SEQ ID NO: 1 in the non-human animal genome with a nucleotide sequence encoding the human IL22 region (e.g., genomic DNA sequence, cDNA sequence, or CDS sequence).

[0214] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding SEQ ID NO: 1 in the non-human animal genome with a portion of exons 2 to 6 of the human IL22 gene.

[0215] Preferably, the non-human animal genome includes replacing parts of exons 1 to 5 of the non-human animal endogenous IL22 gene in the non-human animal genome with parts of exons 2 to 6 of the human IL22 gene.

[0216] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding SEQ ID NO: 1 in the non-human animal genome with the nucleotide sequence encoding SEQ ID NO: 2.

[0217] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding SEQ ID NO: 1 in the non-human animal genome with the nucleotide sequence shown in SEQ ID NO: 5.

[0218] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding SEQ ID NO: 1 in the non-human animal genome with the nucleotide sequence shown in SEQ ID NO: 6.

[0219] Preferably, the expression of human or modified IL22 protein is controlled by endogenous IL22 regulatory elements (e.g., promoters, 5'UTR and / or 3'UTR).

[0220] Preferably, the non-human animal genome is homozygous or heterozygous for the genetically modified endogenous IL22 gene.

[0221] Preferably, the non-human animal is a rodent, preferably a rat or mouse.

[0222] Preferably, the non-human animal genome contains endogenous genes encoding human or modified or wild-type other genes, and preferably, the other genes are selected from at least one of IL22RA1, IL10RB, IL20RA, IL20RB, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3 and OX40.

[0223] Preferably, the non-human animal genome contains an endogenous IL10RB locus encoding a human, modified, or wild-type IL10RB protein.

[0224] In one specific embodiment of the present invention, the non-human animal genome contains a genetically modified endogenous IL10RB locus encoding a modified IL10RB protein, and the non-human animal expresses the modified IL10RB protein.

[0225] Preferably, in the genetically modified endogenous IL10RB locus, the nucleotide sequence encoding the corresponding segment of non-human endogenous IL10RB is replaced with the nucleotide sequence encoding the human IL10RB segment.

[0226] Preferably, the modified IL10RB protein comprises the amino acid sequence shown in SEQ ID NO: 62; or, comprises an amino acid sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in SEQ ID NO: 62; or, comprises an amino acid sequence differing from the amino acid sequence shown in SEQ ID NO: 62 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence similar to that shown in SEQ ID NO: 62, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0227] Preferably, the nucleotide sequence encoding the human IL10RB region includes a portion of the human IL10RB gene; more preferably, it includes a portion of exon 2, all of exon 3, all of exon 4, all of exon 5, and a portion of exon 6 of the human IL10RB gene; even more preferably, it includes portions of exons 2 to 6.

[0228] Preferably, the nucleotide sequence encoding the human IL10RB region comprises the nucleotide sequence shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or a variant thereof; or, comprises a nucleotide sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity to the nucleotide sequence shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or a variant thereof; or, comprises a nucleotide sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity to the nucleotide sequence shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or a variant thereof. The nucleotide sequence shown at positions 33268405-33288111 or its variants differ by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2 or no more than 1 nucleotide; or, a nucleotide sequence containing the nucleotide sequence shown at positions 33268405-33288111 or its variants with NCBI gene accession number NC_000021.9, including substitutions, deletions and / or insertions of one or more nucleotides, wherein the variants include one or more of the following: C>T mutation at position 33276503, G>A mutation at position 33276818, or TTTTGGG>TTG mutation at positions 33277048-33277053;

[0229] Preferably, the mRNA transcribed from the nucleotide sequence encoding the modified IL10RB protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 61; B) the nucleotide sequence having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 61; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 61 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 61, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0230] Preferably, the non-human animal genome contains an endogenous IL20RA locus encoding a human, modified, or wild-type IL20RA protein.

[0231] In one specific embodiment of the present invention, the non-human animal genome contains a genetically modified endogenous IL20RA locus encoding a modified IL20RA protein, and the non-human animal expresses the modified IL20RA protein.

[0232] Preferably, in the genetically modified endogenous IL20RA locus, the nucleotide sequence encoding the corresponding region of non-human endogenous IL20RA is replaced with the nucleotide sequence encoding the human IL20RA region.

[0233] Preferably, the modified IL20RA protein has an amino acid sequence comprising the amino acid sequence shown in SEQ ID NO: 91; or, having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in SEQ ID NO: 91; or, differing from the amino acid sequence shown in SEQ ID NO: 91 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, having an amino acid sequence similar to that shown in SEQ ID NO: 91, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0234] Preferably, the nucleotide sequence encoding the human IL20RA segment includes a portion of the human IL20RA gene; more preferably, it includes a portion of exon 2, all of exon 3, all of exon 4, all of exon 5, and a portion of exon 6 of the human IL20RA gene; even more preferably, it includes portions of exons 2 to 6.

[0235] Preferably, the nucleotide sequence encoding the human IL20RA region comprises the nucleotide sequence shown in positions 137004735-137017092 of NCBI Gene Accession Number NC_000006.12; or, it comprises a nucleotide sequence with an identity of at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99. 8% or at least 99% of the nucleotide sequence; or, a nucleotide sequence that differs from the nucleotide sequence shown in positions 137004735-137017092 of NCBI Gene Accession Number NC_000006.12 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2 or no more than 1 nucleotide; or, a nucleotide sequence that includes substitutions, deletions and / or insertions of one or more nucleotides as shown in the nucleotide sequence shown in positions 137004735-137017092 of NCBI Gene Accession Number NC_000006.12.

[0236] Preferably, the mRNA transcribed from the nucleotide sequence encoding the modified IL20RA protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 90; B) the nucleotide sequence having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 90; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 90 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 90, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0237] Preferably, the non-human animal genome contains an endogenous IL20RB locus encoding a human, modified, or wild-type IL20RB protein.

[0238] In one specific embodiment of the present invention, the non-human animal genome contains a genetically modified endogenous IL20RB locus encoding a modified IL20RB protein, and the non-human animal expresses the modified IL20RB protein.

[0239] Preferably, in the genetically modified endogenous IL20RB locus, the nucleotide sequence encoding the corresponding segment of non-human animal endogenous IL20RB is replaced with the nucleotide sequence encoding the human IL20RB segment.

[0240] Preferably, the modified IL20RB protein has an amino acid sequence comprising the amino acid sequence shown in SEQ ID NO: 121; or, having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in SEQ ID NO: 121; or, differing from the amino acid sequence shown in SEQ ID NO: 121 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, having an amino acid sequence similar to that shown in SEQ ID NO: 121, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0241] Preferably, the nucleotide sequence encoding the human IL20RB region includes a portion of the human IL20RB gene; more preferably, it includes a portion of exon 2, all of exon 3, all of exon 4, all of exon 5, and a portion of exon 6 of the human IL20RB gene; even more preferably, it includes portions of exons 2 to 6.

[0242] Preferably, the nucleotide sequence encoding the human IL20RB region comprises the nucleotide sequence shown in positions 136980507-136995463 of NCBI Gene Accession Number NC_000003.12; or, it comprises a nucleotide sequence with an identity of at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 9 ... 8% or at least 99% of the nucleotide sequence; or, a nucleotide sequence that differs from the nucleotide sequence shown in positions 136980507-136995463 of NCBI Gene Accession Number NC_000003.12 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2 or no more than 1 nucleotide; or, a nucleotide sequence that includes substitutions, deletions and / or insertions of one or more nucleotides as shown in the nucleotide sequence shown in positions 136980507-136995463 of NCBI Gene Accession Number NC_000003.12.

[0243] Preferably, the mRNA transcribed from the nucleotide sequence encoding the modified IL20RB protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 120; B) the nucleotide sequence having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 120; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 120 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 120, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0244] Preferably, the non-human animal genome contains an endogenous IL10 locus encoding a human, modified, or wild-type IL10 protein.

[0245] Preferably, the non-human animal or non-human animal genome contains a genetically modified endogenous IL10 locus encoding a human or modified IL10 protein, and the non-human animal expresses the human or modified IL10 protein.

[0246] Preferably, in the genetically modified endogenous IL10 locus, the nucleotide sequence of the corresponding segment of the non-human animal endogenous IL10 is replaced with the nucleotide sequence encoding the human IL10 segment.

[0247] Preferably, the nucleotide sequence encoding the human IL10 region comprises a portion of exon 1, all of exon 2, all of exon 3, all of exon 4, and a portion of exon 5 of the human IL10 gene; more preferably, it comprises portions of exons 1 to 5. The portion of exon 1 comprises at least 50 bp, for example, at least 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, 160, 165, 170, 180, 190, 200, 220, or 224 bp of continuous nucleotide sequence. The portion of exon 5 contains at least 50 bp, such as at least 50, 60, 70, 80, 90, 93, 100, 200, 300, 400, 500, 600, 700, 800, 900, 1000, 1100, 1120, or 1127 bp of continuous nucleotide sequence.

[0248] Preferably, the non-human animal genome contains an endogenous IL10RA locus encoding a human, modified, or wild-type IL10RA protein.

[0249] Preferably, the non-human animal or the non-human animal genome contains a genetically modified endogenous IL10RA locus encoding a modified IL10RA protein, and the non-human animal expresses the modified IL10RA protein.

[0250] Preferably, in the genetically modified endogenous IL10RA locus, the nucleotide sequence encoding the corresponding segment of non-human animal endogenous IL10RA is replaced with the nucleotide sequence encoding the human IL10RA segment.

[0251] Preferably, the modified IL10RA protein comprises a cytoplasmic domain, a transmembrane domain, and an extracellular domain.

[0252] Preferably, the modified IL10RA protein further comprises a signal peptide.

[0253] Preferably, the IL10RA protein contains a signal peptide that is substantially the same as that of the human IL10RA protein.

[0254] Preferably, the IL10RA protein contains an extracellular domain that is substantially the same as that of the human IL10RA protein.

[0255] Preferably, the IL10RA protein contains a transmembrane domain that is substantially the same as that of the human IL10RA protein.

[0256] Preferably, the IL10RA protein contains a cytoplasmic domain that is substantially the same as that of the human IL10RA protein.

[0257] Preferably, the cytoplasmic domain also includes all or part of the cytoplasmic domain of the non-human animal endogenous IL10RA protein.

[0258] Preferably, the nucleotide sequence encoding the human IL10RA segment includes a portion of exon 1, all of exon 2, all of exon 3, all of exon 4, all of exon 5, and a portion of exon 6 of the human IL10RA gene; more preferably, it includes portions of exons 1 to 6.

[0259] Preferably, the non-human animal genome comprises any one of the following groups:

[0260] 1) Genetically modified endogenous IL22 genes encoding human or modified IL22 proteins and wild-type endogenous IL22RA1 genes;

[0261] 2) Genetically modified endogenous IL22 genes encoding human or modified IL22 proteins, genetically modified endogenous IL22RA1 genes encoding modified IL22RA1 proteins, and wild-type endogenous IL10RB genes.

[0262] 3) Genetically modified endogenous IL22 genes encoding human or modified IL22 proteins, genetically modified endogenous IL22RA1 genes encoding modified IL22RA1 proteins, and genetically modified endogenous IL10RB genes encoding modified IL10RB proteins.

[0263] Preferably, in non-human animals containing the non-human animal genome, the differentiation, development, and distribution of leukocyte subtypes and / or T cell subtypes are not significantly different from those in non-human animals expressing wild-type endogenous IL22, wild-type endogenous IL22RA1, and wild-type endogenous IL10RB.

[0264] Among them, white blood cell subtypes include one or more of B cells, T cells, NK cells, dendritic cells (DCs), granulocytes, macrophages, or monocytes. T cell subtypes include one or more of CD4+ T cells, CD8+ T cells, or Tregs.

[0265] Preferably, the leukocytes or T cells include leukocytes or T cells derived from the spleen, blood, or lymph nodes.

[0266] Preferably, in non-human animals containing the non-human animal genome, the expression levels of CXCL1 and IL6 are not significantly different from those in non-human animals expressing wild-type endogenous IL22, wild-type endogenous IL10RB, and wild-type endogenous IL22RA1.

[0267] In a fourteenth aspect of the invention, a non-human animal is provided, said non-human animal comprising a genetically modified endogenous IL22 gene encoding a human or modified IL22 protein, said non-human animal expressing said human or modified IL22 protein.

[0268] Preferably, the relevant limitations on human or modified IL22 protein and genetically modified endogenous IL22 gene are the same as those in the thirteenth aspect of the present invention.

[0269] Preferably, the non-human animal comprises the non-human animal genome described in aspect thirteen above.

[0270] In a fifteenth aspect, the present invention provides a method for preparing a non-human animal, wherein the non-human animal or the genome of the non-human animal contains a genetically modified endogenous IL22 gene encoding a human or modified IL22 protein, and the non-human animal expresses the human or modified IL22 protein.

[0271] Preferably, the genome of the non-human animal or the non-human animal contains a nucleotide sequence that replaces the nucleotide sequence encoding the corresponding segment of endogenous IL22 in the non-human animal with a nucleotide sequence encoding the human IL22 segment.

[0272] Preferably, the human or modified IL22 protein comprises the amino acid sequence shown in SEQ ID NO: 2; or, comprises an amino acid sequence with at least 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in SEQ ID NO: 2; or, comprises an amino acid sequence differing from the amino acid sequence shown in SEQ ID NO: 2 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence similar to that shown in SEQ ID NO: 2, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0273] Preferably, the nucleotide sequence encoding the human IL22 region includes a portion of exon 2, all of exon 3, all of exon 4, all of exon 5, and a portion of exon 6 of the human IL22 gene.

[0274] Preferably, the nucleotide sequence encoding the human IL22 region comprises the nucleotide sequence shown in SEQ ID NO: 5; or, comprises a nucleotide sequence with an identity of at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the nucleotide sequence shown in SEQ ID NO: 5; or, comprises a nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 5 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or, comprises a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 5, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0275] Preferably, the mRNA transcribed from the nucleotide sequence encoding the human or modified IL22 protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 6; B) the nucleotide sequence having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 6; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 6 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 6, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0276] Preferably, the expression of human or modified IL22 protein is controlled by endogenous IL22 regulatory elements (e.g., promoters, 5'UTR and / or 3'UTR).

[0277] Preferably, the non-human animal or non-human animal genome is homozygous or heterozygous for the genetically modified endogenous IL22 gene.

[0278] Preferably, the non-human animal is a rodent, preferably a rat or mouse.

[0279] Preferably, the genome of the non-human animal or the non-human animal contains an endogenous gene encoding a human or modified other gene. Preferably, the other gene is selected from at least one of IL22RA1, IL10RB, IL20RA, IL20RB, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3, and OX40.

[0280] In a sixteenth aspect, the present invention provides a nucleic acid comprising a nucleotide sequence encoding any of the human or modified IL22 proteins described above. Preferably, the nucleic acid further comprises SEQ ID NO: 5. Preferably, the mRNA transcribed from the nucleic acid comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 6; B) having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 6; C) differing from the nucleotide sequence shown in SEQ ID NO: 6 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) having the nucleotide sequence shown in SEQ ID NO: 6, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0281] In a seventeenth aspect, the present invention provides a humanized IL22 gene, wherein the humanized IL22 gene comprises a portion of the human IL22 gene.

[0282] Preferably, the humanized IL22 gene comprises all or part of exons 1 to 6 of the human IL22 gene. More preferably, the humanized IL22 gene comprises all or part of any one, two, three or more, or consecutive combinations of two or more exons 1 to 6 of the human IL22 gene. More preferably, it comprises all or part of exons 2 to 6. Even more preferably, the humanized IL22 gene comprises a portion of exon 2, all of exons 3 to 5, and a portion of exon 6 of the human IL22 gene, preferably also comprising intron 2 and / or intron 5, and more preferably comprising any intron between exons 2 and 6. In one specific embodiment of the present invention, the humanized IL22 gene comprises portions of exons 2 to 6 of the human IL22 gene, wherein the portion of exon 2 comprises at least 50 bp of continuous nucleotide sequence, for example, at least 50, 70, 90, 100, 150, 170, 180, 185, 186, 187, 188, 189, 190, 200, and 232 bp of nucleotide sequence; preferably, the portion of exon 2 comprises from the start codon to the last nucleotide of exon 2, and the portion of exon 6 comprises at least 20 bp of continuous nucleotide sequence, for example, at least 20, 30, 40, 50, 70, 75, 76, 77, 78, 79, 80, 100, 200, 500, 600, and 635 bp of nucleotide sequence; preferably, the portion of exon 6 comprises from the first nucleotide of exon 6 to the stop codon.

[0283] In one specific embodiment of the present invention, the humanized IL22 gene comprises the nucleotide sequence shown in SEQ ID NO: 5; or, comprises a nucleotide sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 5; or, comprises a nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 5 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or, comprises a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 5, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0284] Preferably, the humanized IL22 gene comprises all or part of the nucleotide sequence encoding the human IL22 protein. More preferably, the humanized IL22 gene comprises the nucleotide sequence encoding SEQ ID NO: 2; or, comprises a nucleotide sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity to the nucleotide sequence encoding SEQ ID NO: 2; or, comprises a nucleotide sequence differing from the amino acid sequence encoding SEQ ID NO: 2 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises a nucleotide sequence having the nucleotide sequence encoding SEQ ID NO: 2, including nucleotide sequences with substitutions, deletions, and / or insertions of one or more nucleotides.

[0285] Preferably, the humanized IL22 gene further comprises all or part of the non-human animal IL22 gene. More preferably, it comprises all or part of exons 1 to 5 of the non-human animal IL22 gene; more preferably, it comprises a portion of exon 1 and / or exon 5 of the non-human animal IL22 gene; and even more preferably, it comprises the 5'UTR and / or 3'UTR of the non-human animal IL22 gene.

[0286] In one specific embodiment of the present invention, the nucleotide sequence of the humanized IL22 gene comprises any one of the following groups: A) the transcribed mRNA is the nucleotide sequence shown in SEQ ID NO: 6; B) the transcribed mRNA is at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identical to the nucleotide sequence shown in SEQ ID NO: 6; C) the transcribed mRNA differs from the nucleotide sequence shown in SEQ ID NO: 6 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) the transcribed mRNA has the nucleotide sequence shown in SEQ ID NO: 6, including nucleotide sequences with substitutions, deletions, and / or insertions of one or more nucleotides.

[0287] Preferably, the humanized IL22 gene further includes a specific inducer or repressor. More preferably, the specific inducer or repressor can be a conventionally induced or repressible substance. In one specific embodiment of the present invention, the specific inducer is selected from the tetracycline system (Tet-Off System / Tet-On System) or the tamoxifen system.

[0288] Preferably, the non-human animal can be selected from rodents, pigs, rabbits, monkeys, or any other non-human animal that can be genetically edited to produce humanized genes. Further limitations on the type of non-human animal are the same as in the sixth aspect of this invention.

[0289] In an eighteenth aspect, the present invention provides a humanized IL22 protein, wherein the humanized IL22 protein is encoded by the aforementioned humanized IL22 gene.

[0290] Preferably, the humanized IL22 protein comprises all or part of the human IL22 protein.

[0291] Preferably, the humanized IL22 protein comprises all or part of the amino acid sequence encoded by exons 1 to 6 of the human IL22 gene. More preferably, it comprises all or part of the amino acid sequence encoded by any one, two, three or more, or a combination of two or more consecutive exons 1 to 6. More preferably, it comprises all or part of the amino acid sequence encoded by exons 2 to 6. Even more preferably, it comprises a portion of exon 2, all of exons 3 to 5, and a portion of exon 6. More preferably, it comprises a portion of exons 2 to 6, wherein the portion of exon 2 comprises at least 50 bp of continuous nucleotide sequence, for example, at least 50, 70, 90, 100, 150, 170, 180, 185, 186, 187, 188, 189, 190, 200, and 232 bp. The nucleotide sequence; preferably, the exon 2 portion includes the sequence from the start codon to the last nucleotide of exon 2, and the exon 6 portion includes at least 20 bp of continuous nucleotide sequence, for example, at least 20, 30, 40, 50, 70, 75, 76, 77, 78, 79, 80, 100, 200, 500, 600, and 635 bp of nucleotide sequence; preferably, the exon 6 portion includes the sequence from the first nucleotide of exon 6 to the stop codon.

[0292] Preferably, the humanized IL22 protein comprises the amino acid sequence shown in SEQ ID NO: 2; or, comprises an amino acid sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in SEQ ID NO: 2; or, comprises an amino acid sequence differing from the amino acid sequence shown in SEQ ID NO: 2 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence similar to that shown in SEQ ID NO: 2, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0293] Preferably, the non-human animal can be selected from rodents, pigs, rabbits, monkeys, or any other non-human animal that can be genetically edited to produce humanized genes. Further limitations on the type of non-human animal are the same as in the sixth aspect of this invention.

[0294] A nineteenth aspect of the present invention provides a targeting vector comprising a donor nucleotide sequence comprising any one of the following groups: A) a nucleotide sequence encoding a humanized IL22 protein; B) a nucleotide sequence of a humanized IL22 gene; C) a portion of a human IL22 gene, preferably comprising all or a portion of exons 1 to 6 of the human IL22 gene. Preferably, it comprises all or a portion of any one, two, three or more, or a combination of two or more consecutive exons 1 to 6. Further preferred are those containing all or part of exons 2 to 6 of the human IL22 gene; even more preferred are those containing a portion of exon 2, all of exons 3 to 5, and a portion of exon 6; even more preferred are those containing a portion of exons 2 to 6, wherein the exon 2 portion contains at least a 50 bp continuous nucleotide sequence, for example, at least 50, 70, 90, 100, 150, 170, 180, 185, 186, 187, 188, 189, 190, 20... The nucleotide sequence is 0 or 232 bp; preferably, the exon 2 portion includes the sequence from the start codon to the last nucleotide of exon 2, and the exon 6 portion includes at least 20 bp of continuous nucleotide sequence, for example, at least 20, 30, 40, 50, 70, 75, 76, 77, 78, 79, 80, 100, 200, 500, 600, or 635 bp; preferably, the exon 6 portion includes the sequence from the first nucleotide of exon 6 to the stop codon. Further preferably, the donor nucleotide sequence comprises the nucleotide sequence shown in SEQ ID NO: 5; or, comprises a nucleotide sequence having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 5; or, comprises a nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 5 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or, comprises a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 5, including substitutions, deletions, and / or insertions of one or more nucleotides.Alternatively, D) a nucleotide sequence encoding all or part of the human IL22 protein, preferably comprising the nucleotide sequence encoding SEQ ID NO: 2, or comprising a nucleotide sequence having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence encoding SEQ ID NO: 2; or comprising a nucleotide sequence differing from the nucleotide sequence encoding SEQ ID NO: 2 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or comprising a nucleotide sequence having the nucleotide sequence encoding SEQ ID NO: 2, including nucleotide sequences with substitutions, deletions, and / or insertions of one or more nucleotides.

[0295] Preferably, the targeting vector further comprises a 5' arm (5' homologous arm) and / or a 3' arm (3' homologous arm).

[0296] The 5' arm is a DNA fragment homologous to the 5' end of the conversion region to be altered, selected from 100-10000 nucleotides in length from the genomic DNA of the non-human animal IL22 gene. Preferably, the 5' arm has at least 90% homology with the nucleotide in NCBI accession number NC_000076.7. More preferably, the 5' arm sequence comprises SEQ ID NO: 3.

[0297] The 3' arm is a DNA fragment homologous to the 3' end of the conversion region to be modified, and is selected from 100-10000 nucleotides in length from the genomic DNA of the IL22 gene in non-human animals; preferably, the 3' arm has at least 90% homology with the nucleotide in NCBI accession number NC_000076.7; more preferably, the 3' arm sequence contains the sequence shown in SEQ ID NO: 4.

[0298] Preferably, the targeting vector further comprises SEQ ID NO: 7 and / or 8.

[0299] Preferably, the conversion region to be altered is located on exons 1 to 5 of the non-human animal IL22 gene, and more preferably on the start codon to the stop codon of the non-human animal IL22 gene.

[0300] Preferably, the targeting vector further comprises a marker gene, a resistance gene for positive clone selection, and / or a specific recombination system. The relevant limitations regarding the marker gene, the resistance gene for positive clone selection, and / or the specific recombination system are the same as in aspect eight of the present invention.

[0301] A twentieth aspect of the invention provides a cell comprising the targeting vector described in the nineteenth aspect.

[0302] In a twenty-first aspect, the present invention provides the application of the targeting vector described in the nineteenth aspect and / or the cells described in the twentieth aspect in IL22 gene editing, preferably, the application including but not limited to knockout, insertion or replacement.

[0303] In a twenty-second aspect, the present invention provides a non-human animal with a humanized IL22 gene, wherein the non-human animal expresses human or humanized IL22 protein.

[0304] Preferably, the humanized IL22 protein described above is expressed in the non-human animal.

[0305] Preferably, the expression of endogenous IL22 protein in the non-human animal is reduced or absent.

[0306] Preferably, the non-human animal genome contains a portion of the human IL22 gene or a humanized IL22 gene, more preferably it contains the aforementioned humanized IL22 gene.

[0307] Preferably, a portion of the human IL22 gene or the nucleotide sequence of the humanized IL22 gene is operatively linked to an endogenous regulatory element at an endogenous IL22 gene locus in at least one chromosome.

[0308] Preferably, the non-human animal is constructed by introducing a donor nucleotide sequence into the non-human animal IL22 locus. Preferably, the limitation of the donor nucleotide sequence is the same as in the nineteenth aspect of the present invention.

[0309] Preferably, the non-human animal is constructed using the aforementioned targeting vector.

[0310] According to some embodiments of the present invention, the non-human animal further comprises other gene modifications, said other genes being selected from at least one of IL22RA1, IL10RB, IL20RA, IL20RB, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3, and OX40.

[0311] Preferably, the IL22RA1 gene is a humanized IL22RA1 gene; more preferably, the humanized IL22RA1 gene is any of the humanized IL22RA1 genes described in this invention.

[0312] Preferably, the IL10RB gene is a humanized IL10RB gene; more preferably, the humanized IL10RB gene is any of the humanized IL10RB genes described in this invention.

[0313] Preferably, the IL20RA gene is a humanized IL20RA gene; more preferably, the humanized IL20RA gene is any of the humanized IL20RA genes described in this invention.

[0314] Preferably, the IL20RB gene is a humanized IL20RB gene; more preferably, the humanized IL20RB gene is any of the humanized IL20RB genes described in this invention.

[0315] According to some embodiments of the present invention, the partial or humanized IL22 gene of the human IL22 gene and / or the other genes are homozygous or heterozygous for the endogenously replaced locus.

[0316] Preferably, the humanized IL22 gene further includes a specific inducer or repressor; more preferably, the specific inducer or repressor can be a conventionally induced or repressible substance. In one specific embodiment of the present invention, the specific inducer is selected from the tetracycline system (Tet-Off System / Tet-On System) or the tamoxifen system.

[0317] Preferably, the non-human animal can be selected from rodents, pigs, rabbits, monkeys, or any other non-human animal that can be genetically edited to produce humanized genes. Further limitations on the type of non-human animal are the same as in the sixth aspect of this invention.

[0318] In a twenty-third aspect, the present invention provides a method for constructing a non-human animal with humanized IL22 gene, wherein the non-human animal expresses human or humanized IL22 protein.

[0319] Preferably, the humanized IL22 protein described above is expressed in the non-human animal.

[0320] Preferably, the genome of at least one cell of the non-human animal contains a portion of the human IL22 gene or a humanized IL22 gene, more preferably the aforementioned humanized IL22 gene.

[0321] Preferably, the non-human animal is a non-human animal with the IL22 gene humanized as described above.

[0322] Preferably, the construction method includes introducing a donor nucleotide sequence into the IL22 gene locus of a non-human animal. Preferably, the limitation of the donor nucleotide sequence is the same as that in the nineteenth aspect of the present invention.

[0323] Preferably, the human or humanized IL22 gene is operatively linked to at least one endogenous IL22 regulatory element on a chromosome.

[0324] Preferably, the partial or humanized human IL22 gene is regulated in non-human animals by a regulatory element. More preferably, the regulatory element can be endogenous or exogenous.

[0325] Preferably, the control elements include, but are not limited to, a promoter, a 5'UTR, and / or a 3'UTR.

[0326] In one specific embodiment of the present invention, the endogenous regulatory element is derived from the non-human animal IL22 gene. The exogenous regulatory element is derived from the human IL22 gene.

[0327] Preferably, the insertion site is located after the endogenous regulatory element of the IL22 gene.

[0328] Preferably, the introduction is a substitution, insertion, or transgenesis. Optionally, the introduction of the non-human animal IL22 gene locus involves replacing the corresponding region in the non-human animal. More preferably, all or part of exons 1 to 5 of the non-human animal IL22 gene are replaced. More preferably, a portion of exon 1, all of exons 2 to 4, and a portion of exon 5 of the non-human animal IL22 gene are replaced. Even more preferably, a portion of exons 1 to 5 of the non-human animal IL22 gene are replaced. In one specific embodiment of the invention, the nucleotide sequence from the start codon to the stop codon of the non-human animal IL22 gene is replaced.

[0329] Preferably, the nucleotide sequence encoding the amino acid shown in SEQ ID NO:1 is replaced.

[0330] Preferably, the non-human animal used to construct the humanized IL22 gene is constructed using gene editing technology, wherein the gene editing technology includes gene targeting technology using embryonic stem cells, CRISPR / Cas9 technology, zinc finger nuclease technology, transcription activator-like effector nuclease technology, homing endonuclease or other molecular biology techniques.

[0331] In one specific embodiment of the present invention, the construction method includes modifying the coding frame of a non-human animal IL22 gene by inserting a nucleotide sequence encoding a human or humanized IL22 protein, or a portion of the human IL22 gene or a nucleotide sequence of the humanized IL22 gene, after the endogenous regulatory element of the non-human animal IL22 gene. The modified coding frame of the non-human animal IL22 gene can be a knockout of the functional region of the non-human animal IL22 gene or the insertion of a sequence, causing the non-human animal IL22 protein to be unexpressed, underexpressed, or non-functional. More preferably, the modified coding frame of the non-human animal IL22 gene can be all or part of exons 1 to 5 of the non-human animal IL22 gene, preferably a portion of exon 1, all of exons 2 to 4, and a portion of exon 5, and even more preferably a portion of exons 1 to 5 of the non-human animal IL22 gene. In one specific embodiment of the present invention, the coding frame of the modified non-human animal IL22 gene can be a nucleotide sequence from the start codon to the stop codon of the non-human animal IL22 gene.

[0332] Preferably, the above-mentioned targeting vector is used to construct non-human animals.

[0333] In one specific embodiment of the present invention, the construction method includes introducing the above-mentioned targeting vector into non-human animal cells (preferably embryonic stem cells), screening out the correct positive clone cells and introducing them into the isolated blastocyst, culturing the blastocyst, and then transplanting the cultured blastocyst into the oviduct of a female non-human animal, allowing it to develop, and identifying and screening to obtain non-human animals with humanized IL22 gene.

[0334] According to some embodiments of the present invention, the construction method further includes: mating non-human animals with humanized IL22 genes with other genetically modified non-human animals, in vitro fertilization or direct gene editing, and screening to obtain multi-gene-modified non-human animals.

[0335] Preferably, the other genes are non-human animals modified with at least one of the following genes: IL22RA1, IL10RB, IL20RA, IL20RB, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3, and OX40.

[0336] Preferably, the non-human animal also expresses at least one of the following human or humanized proteins: IL22RA1, IL10RB, IL20RA, IL20RB, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3, and OX40.

[0337] Preferably, the IL22RA1 gene is a humanized IL22RA1 gene; more preferably, the humanized IL22RA1 gene is any of the humanized IL22RA1 genes described in this invention.

[0338] Preferably, the IL10RB gene is a humanized IL10RB gene; more preferably, the humanized IL10RB gene is any of the humanized IL10RB genes described in this invention.

[0339] Preferably, the IL20RA gene is a humanized IL20RA gene; more preferably, the humanized IL20RA gene is any of the humanized IL20RA genes described in this invention.

[0340] Preferably, the IL20RB gene is a humanized IL20RB gene; more preferably, the humanized IL20RB gene is any of the humanized IL20RB genes described in this invention.

[0341] Preferably, the IL22RA1 protein is a humanized IL22RA1 protein; more preferably, the humanized IL22RA1 protein is any of the humanized IL22RA1 proteins described in this invention.

[0342] Preferably, the IL10RB protein is a humanized IL10RB protein, and more preferably, the humanized IL10RB protein is any of the humanized IL10RB proteins described in this invention.

[0343] Preferably, the IL20RA protein is a humanized IL20RA protein; more preferably, the humanized IL20RA protein is any of the humanized IL20RA proteins described in this invention.

[0344] Preferably, the IL20RB protein is a humanized IL20RB protein; more preferably, the humanized IL20RB protein is any of the humanized IL20RB proteins described in this invention.

[0345] Preferably, in the genome of the multi-gene modified non-human animal, each of the modified genes is homozygous or heterozygous for the endogenous replaced locus.

[0346] Preferably, the non-human animal can be selected from rodents, pigs, rabbits, monkeys, or any other non-human animal that can be genetically edited to produce humanized genes. Further limitations on the type of non-human animal are the same as in the sixth aspect of this invention.

[0347] In a twenty-fourth aspect, the present invention provides a non-human animal genome, wherein the non-human animal or the non-human animal genome comprises a genetically modified endogenous IL10RB gene encoding a modified IL10RB protein, and the non-human animal expresses the modified IL10RB protein.

[0348] Preferably, the non-human animal or non-human animal genome contains a nucleotide sequence encoding the corresponding segment of the non-human animal endogenous IL10RB, which is replaced by a nucleotide sequence encoding the human IL10RB segment.

[0349] Preferably, the modified IL10RB protein comprises a cytoplasmic domain, a transmembrane domain, and an extracellular domain;

[0350] Preferably, the modified IL10RB protein contains an extracellular domain substantially the same as that of the human IL10RB protein.

[0351] Preferably, all or part of the extracellular domain comprises the amino acid sequence shown at positions 20-220 or 21-218 of SEQ ID NO: 57; or, comprises an amino acid sequence with an identity of at least 77%, 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown at positions 20-220 or 21-218 of SEQ ID NO: 57; or, comprises an amino acid sequence differing from the amino acid sequence shown at positions 20-220 or 21-218 of SEQ ID NO: 57 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with an amino acid sequence shown at positions 20-220 or 21-218 of SEQ ID NO: 57, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0352] Preferably, the extracellular domain comprises 1-5 amino acid residues (e.g., 1, 2, 3, 4, or 5) at the N-terminus and / or C-terminus of the extracellular domain of the non-human endogenous IL10RB protein, further comprising the amino acid sequence shown at position 22 or positions 221-222 of SEQ ID NO: 56; or, comprising an amino acid sequence with an identity of at least 77%, 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown at position 22 or positions 221-222 of SEQ ID NO: 56; or, comprising an amino acid sequence with an identity of at least 10, 9, 8, 7, 6, 5, 4, 3, 2, or at least 1 amino acid ... The amino acid sequence shown at position 22 or positions 221-222 of NO:56, including the substitution, deletion and / or insertion of one or more amino acid residues.

[0353] Preferably, the non-human animal is a rodent, and more preferably a rat or mouse.

[0354] Preferably, the modified IL10RB protein contains a cytoplasmic domain that is substantially the same as that of the non-human animal endogenous IL10RB protein.

[0355] Preferably, the amino acid sequence of the cytoplasmic domain comprises the amino acid sequence shown in positions 244-351 of SEQ ID NO: 56; or, comprises an amino acid sequence with an identity of at least 77%, 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown in positions 244-351 of SEQ ID NO: 56; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 244-351 of SEQ ID NO: 56 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with an amino acid sequence shown in positions 244-351 of SEQ ID NO: 56, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0356] Preferably, the modified IL10RB protein contains a transmembrane domain that is substantially the same as that of the non-human endogenous IL10RB protein.

[0357] Preferably, the amino acid sequence of the transmembrane domain comprises the amino acid sequence shown in positions 223-243 of SEQ ID NO: 56; or, comprises an amino acid sequence with an identity of at least 77%, 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown in positions 223-243 of SEQ ID NO: 56; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 223-243 of SEQ ID NO: 56 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with an amino acid sequence shown in positions 223-243 of SEQ ID NO: 56, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0358] Preferably, the modified IL10RB protein has an amino acid sequence comprising the amino acid sequence shown in SEQ ID NO: 62; or, having at least 77%, 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in SEQ ID NO: 62; or, differing from the amino acid sequence shown in SEQ ID NO: 62 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, having an amino acid sequence showing substitutions, deletions, and / or insertions of one or more amino acid residues as shown in the amino acid sequence shown in SEQ ID NO: 62.

[0359] Preferably, the nucleotide sequence of the replaced region encoding the corresponding region of non-human endogenous IL10RB comprises all or part of the nucleotide sequence encoding the extracellular domain of the non-human endogenous IL10RB protein, more preferably comprising the nucleotide sequence encoding positions 23-220 or 22-222 of SEQ ID NO: 56, or comprising a nucleotide sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence encoding positions 23-220 or 22-222 of SEQ ID NO: 56; or comprising a nucleotide sequence with a difference of no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide from ... The nucleotide sequence shown at positions 23-220 or 22-222 of NO: 56 includes a nucleotide sequence with substitutions, deletions, and / or insertions of one or more nucleotides. More preferably, it includes portions of exons 2 to 6 of the non-human animal endogenous IL10RB sequence.

[0360] Preferably, the nucleotide sequence encoding the human IL10RB region comprises a portion of the human IL10RB gene; more preferably, it comprises all or part of exons 2 to 6 of the human IL10RB gene, and even more preferably, it comprises a portion of exon 2, all of exon 3, all of exon 4, all of exon 5, and a portion of exon 6. Preferably, the nucleotide sequence encoding the human IL10RB region comprises portions of exons 2 to 6. The exon 2 portion contains at least 50 bp of nucleotide sequence, for example, at least 50, 60, 70, 80, 90, 100, 110, 113, 120, or 124 bp of nucleotide sequence; the exon 6 portion contains at least 5 bp of nucleotide sequence, for example, at least 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, or 158 bp of nucleotide sequence.

[0361] Preferably, the nucleotide sequence encoding the human IL10RB region comprises the nucleotide sequence shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or a variant thereof, or comprises a nucleotide sequence showing at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, or 96% identity with the nucleotide sequence shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or a variant thereof. 97%, 98%, or at least 99%; or, containing a nucleotide sequence that differs from the nucleotide sequence shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or a variant thereof by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or, containing a nucleotide sequence that, including substitutions, deletions, and / or insertions of one or more nucleotides, is shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or a variant thereof.

[0362] The variants include one or more of the following: C>T mutation at position 33276503, G>A mutation at position 33276818, or TTTGTGG>TTG mutation at positions 33277048-33277053.

[0363] Preferably, the mRNA transcribed from the nucleotide sequence encoding the modified IL10RB protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 61; B) the nucleotide sequence having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 61; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 61 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 61, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0364] Preferably, the non-human animal genome includes replacing all or part of the nucleotide sequence encoding the extracellular domain of the endogenous IL10RB protein in the non-human animal genome with a nucleotide sequence encoding an extracellular domain substantially identical to that of the human IL10RB protein.

[0365] Preferably, the non-human animal genome includes replacing all or part of the nucleotide sequence encoding the extracellular domain of the endogenous IL10RB protein in the non-human animal genome with a nucleotide sequence encoding the human IL10RB region.

[0366] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding positions 23-220 or 22-222 of SEQ ID NO: 56 in the non-human animal genome with a nucleotide sequence encoding the human IL10RB region (e.g., genomic DNA sequence, cDNA sequence, or CDS sequence).

[0367] Preferably, the non-human animal genome includes replacing parts of exons 2 to 6 of the non-human animal endogenous IL10RB gene in the non-human animal genome with parts of exons 2 to 6 of the human IL10RB gene.

[0368] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding SEQ ID NO: 56 from position 23 to 220 in the non-human animal genome with the nucleotide sequence encoding positions 21-218 of SEQ ID NO: 57.

[0369] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding SEQ ID NO: 56 from position 22 to 222 in the non-human animal genome with the nucleotide sequence encoding positions 20-220 of SEQ ID NO: 57.

[0370] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding positions 22-222 of SEQ ID NO: 56 in the non-human animal genome with the nucleotide sequence encoding SEQ ID NO: 62.

[0371] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding positions 23-220 of SEQ ID NO: 56 in the non-human animal genome with the nucleotide sequence shown at positions 33268405-33288111 of NCBI accession number NC_000021.9 or a variant thereof. The variant includes one or more of the following: a C>T mutation at position 33276503, a G>A mutation at position 33276818, or a TTTTGGG>TTG mutation at positions 33277048-33277053.

[0372] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding SEQ ID NO: 56 in the non-human animal genome with the nucleotide sequence of SEQ ID NO: 61.

[0373] Preferably, the expression of the modified IL10RB protein is controlled by endogenous IL10RB regulatory elements (e.g., promoter, 5'UTR and / or 3'UTR).

[0374] Preferably, the non-human animal genome is homozygous or heterozygous for the genetically modified endogenous IL10RB gene.

[0375] Preferably, the non-human animal genome contains endogenous genes encoding human or modified or wild-type other genes, and preferably, the other genes are selected from at least one of IL22, IL22RA, IL20RA, IL20RB, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3 and OX40.

[0376] Preferably, the non-human animal genome contains an endogenous IL10 locus encoding a human, modified, or wild-type IL10 protein.

[0377] Preferably, the non-human animal or non-human animal genome contains a genetically modified endogenous IL10 locus encoding a human or modified IL10 protein, and the non-human animal expresses the human or modified IL10 protein.

[0378] Preferably, in the genetically modified endogenous IL10 locus, the nucleotide sequence of the corresponding segment of the non-human animal endogenous IL10 is replaced with the nucleotide sequence encoding the human IL10 segment.

[0379] Preferably, the nucleotide sequence encoding the human IL10 region comprises a portion of exon 1, all of exon 2, all of exon 3, all of exon 4, and a portion of exon 5 of the human IL10 gene; more preferably, it comprises portions of exons 1 to 5. The portion of exon 1 comprises at least 50 bp, for example, at least 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, 160, 165, 170, 180, 190, 200, 220, or 224 bp of continuous nucleotide sequence. The portion of exon 5 contains at least 50 bp, such as at least 50, 60, 70, 80, 90, 93, 100, 200, 300, 400, 500, 600, 700, 800, 900, 1000, 1100, 1120, or 1127 bp of continuous nucleotide sequence.

[0380] Preferably, the non-human animal genome contains an endogenous IL10RA locus encoding a human, modified, or wild-type IL10RA protein.

[0381] Preferably, the non-human animal or the non-human animal genome contains a genetically modified endogenous IL10RA locus encoding a modified IL10RA protein, and the non-human animal expresses the modified IL10RA protein.

[0382] Preferably, in the genetically modified endogenous IL10RA locus, the nucleotide sequence encoding the corresponding segment of non-human animal endogenous IL10RA is replaced with the nucleotide sequence encoding the human IL10RA segment.

[0383] Preferably, the modified IL10RA protein comprises a cytoplasmic domain, a transmembrane domain, and an extracellular domain.

[0384] Preferably, the modified IL10RA protein further comprises a signal peptide.

[0385] Preferably, the IL10RA protein contains a signal peptide that is substantially the same as that of the human IL10RA protein.

[0386] Preferably, the IL10RA protein contains an extracellular domain that is substantially the same as that of the human IL10RA protein.

[0387] Preferably, the IL10RA protein contains a transmembrane domain that is substantially the same as that of the human IL10RA protein.

[0388] Preferably, the IL10RA protein contains a cytoplasmic domain that is substantially the same as that of the human IL10RA protein.

[0389] Preferably, the cytoplasmic domain also includes all or part of the cytoplasmic domain of the non-human animal endogenous IL10RA protein.

[0390] Preferably, the nucleotide sequence encoding the human IL10RA segment includes a portion of exon 1, all of exon 2, all of exon 3, all of exon 4, all of exon 5, and a portion of exon 6 of the human IL10RA gene; more preferably, it includes portions of exons 1 to 6.

[0391] Preferably, the non-human animal genome comprises any one of the following groups:

[0392] 1) Genetically modified endogenous IL10RB gene, wild-type endogenous IL22 gene, and wild-type endogenous IL22RA1 gene encoding modified IL10RB protein;

[0393] 2) Genetically modified endogenous IL10RB genes encoding modified IL10RB proteins, genetically modified endogenous IL22 genes encoding human or modified IL22 proteins, and wild-type endogenous IL22RA1 genes;

[0394] 3) Genetically modified endogenous IL10RB gene encoding modified IL10RB protein, genetically modified endogenous IL22RA1 gene encoding modified IL22RA1 protein, and wild-type endogenous IL22 gene;

[0395] 4) Genetically modified endogenous IL10RB genes encoding modified IL10RB protein, genetically modified endogenous IL22RA1 genes encoding modified IL22RA1 protein, and genetically modified endogenous IL22 genes encoding human or modified IL22 protein.

[0396] 5) Genetically modified endogenous IL10RB gene, wild-type endogenous IL20RA gene, and wild-type endogenous IL26 gene encoding modified IL10RB protein;

[0397] 6) Genetically modified endogenous IL10RB genes encoding modified IL10RB protein, genetically modified endogenous IL20RA genes encoding human or modified IL20RA protein, and wild-type endogenous IL26 genes.

[0398] 7) Genetically modified endogenous IL10RB gene encoding modified IL10RB protein, genetically modified endogenous IL26 gene encoding modified IL26 protein, and wild-type endogenous IL20RA gene;

[0399] 8) Genetically modified endogenous IL10RB gene encoding modified IL10RB protein, genetically modified endogenous IL20RA gene encoding modified IL20RA protein, and genetically modified endogenous IL26 gene encoding human or modified IL26 protein.

[0400] 9) Genetically modified endogenous IL10RB gene, wild-type endogenous IL10RA gene and wild-type endogenous IL10RA gene encoding modified IL10RB protein;

[0401] 10) Genetically modified endogenous IL10RB genes encoding modified IL10RB proteins, genetically modified endogenous IL10RA genes encoding human or modified IL20RA proteins, and wild-type endogenous IL10 genes.

[0402] 11) Genetically modified endogenous IL10RB gene encoding modified IL10RB protein, genetically modified endogenous IL10 gene encoding modified IL26 protein, and wild-type endogenous IL10RA gene;

[0403] 12) Genetically modified endogenous IL10RB genes encoding modified IL10RB proteins, genetically modified endogenous IL10RA genes encoding modified IL10RA proteins, and genetically modified endogenous IL10 genes encoding human or modified IL10 proteins.

[0404] Preferably, in non-human animals containing the non-human animal genome, the differentiation, development, and distribution of leukocyte subtypes and / or T cell subtypes are not significantly different from those in non-human animals expressing wild-type endogenous IL22, wild-type endogenous IL22RA1, and wild-type endogenous IL10RB.

[0405] Among them, white blood cell subtypes include one or more of B cells, T cells, NK cells, dendritic cells (DCs), granulocytes, macrophages, or monocytes. T cell subtypes include one or more of CD4+ T cells, CD8+ T cells, or Tregs.

[0406] Preferably, the leukocytes or T cells include leukocytes or T cells derived from the spleen, blood, or lymph nodes.

[0407] Preferably, in non-human animals containing the non-human animal genome, the expression levels of CXCL1 and IL6 are not significantly different from those in non-human animals expressing wild-type endogenous IL22, wild-type endogenous IL10RB, and wild-type endogenous IL22RA1.

[0408] In a twenty-fifth aspect, the present invention provides a non-human animal comprising a genetically modified endogenous IL10RB gene encoding a modified IL10RB protein, wherein the non-human animal expresses the modified IL10RB protein.

[0409] Preferably, the relevant limitations on the modified IL10RB protein and the genetically modified endogenous IL10RB gene are the same as those in aspect twenty-four of the present invention.

[0410] Preferably, the non-human animal comprises the non-human animal genome described in aspect twenty-four.

[0411] In a twenty-sixth aspect, the present invention provides a method for preparing a non-human animal, said non-human animal comprising a genetically modified endogenous IL10RB gene encoding a modified IL10RB protein, said non-human animal expressing said modified IL10RB protein.

[0412] Preferably, the genome of the non-human animal or the non-human animal contains a nucleotide sequence that replaces the nucleotide sequence encoding the corresponding segment of the non-human animal's endogenous IL10RB with a nucleotide sequence encoding the human IL10RB segment.

[0413] Preferably, the modified IL10RB protein comprises a cytoplasmic domain, a transmembrane domain, and an extracellular domain.

[0414] Preferably, the modified IL10RB protein contains an extracellular domain substantially the same as that of the human IL10RB protein.

[0415] Preferably, all or part of the extracellular domain comprises the amino acid sequence shown at positions 20-220 or 21-218 of SEQ ID NO: 57; or, comprises an amino acid sequence with an identity of at least 77%, 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown at positions 20-220 or 21-218 of SEQ ID NO: 57; or, comprises an amino acid sequence differing from the amino acid sequence shown at positions 20-220 or 21-218 of SEQ ID NO: 57 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with an amino acid sequence shown at positions 20-220 or 21-218 of SEQ ID NO: 57, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0416] Preferably, the non-human animal is a rodent, and more preferably a rat or mouse.

[0417] Preferably, the modified IL10RB protein contains a cytoplasmic domain that is substantially the same as that of the non-human animal endogenous IL10RB protein.

[0418] Preferably, the amino acid sequence of the cytoplasmic domain comprises the amino acid sequence shown in positions 244-351 of SEQ ID NO: 56; or, comprises an amino acid sequence with an identity of at least 77%, 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown in positions 244-351 of SEQ ID NO: 56; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 244-351 of SEQ ID NO: 56 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with an amino acid sequence shown in positions 244-351 of SEQ ID NO: 56, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0419] Preferably, the modified IL10RB protein contains a transmembrane domain that is substantially the same as that of the non-human endogenous IL10RB protein.

[0420] Preferably, the amino acid sequence of the transmembrane domain comprises the amino acid sequence shown in positions 223-243 of SEQ ID NO: 56; or, comprises an amino acid sequence with an identity of at least 77%, 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown in positions 223-243 of SEQ ID NO: 56; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 223-243 of SEQ ID NO: 56 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with an amino acid sequence shown in positions 223-243 of SEQ ID NO: 56, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0421] Preferably, the modified IL10RB protein has an amino acid sequence comprising the amino acid sequence shown in SEQ ID NO: 62; or, having at least 77%, 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in SEQ ID NO: 62; or, differing from the amino acid sequence shown in SEQ ID NO: 62 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, having an amino acid sequence showing substitutions, deletions, and / or insertions of one or more amino acid residues as shown in the amino acid sequence shown in SEQ ID NO: 62.

[0422] Preferably, the nucleotide sequence encoding the human IL10RB region includes a portion of exon 2, all of exon 3, all of exon 4, all of exon 5, and a portion of exon 6 of the human IL10RB gene.

[0423] Preferably, the nucleotide sequence encoding the human IL10RB region comprises the nucleotide sequence shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or a variant thereof, or comprises a nucleotide sequence showing at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, or 96% identity with the nucleotide sequence shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or a variant thereof. 97%, 98%, or at least 99%; or, containing a nucleotide sequence that differs from the nucleotide sequence shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or a variant thereof by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or, containing a nucleotide sequence that includes substitutions, deletions, and / or insertions of one or more nucleotides as shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or a variant thereof.

[0424] The variants include one or more of the following: C>T mutation at position 33276503, G>A mutation at position 33276818, or TTTGTGG>TTG mutation at positions 33277048-33277053.

[0425] Preferably, the mRNA transcribed from the nucleotide sequence encoding the modified IL10RB protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 61; B) the nucleotide sequence having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 61; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 61 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 61, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0426] Preferably, the expression of the modified IL10RB protein is controlled by endogenous IL10RB regulatory elements (e.g., promoter, 5'UTR and / or 3'UTR).

[0427] Preferably, the non-human animal genome is homozygous or heterozygous for the genetically modified endogenous IL10RB gene.

[0428] Preferably, the genome of the non-human animal or the non-human animal contains an endogenous gene encoding a human or modified or wild-type other gene. Preferably, the other gene is selected from at least one of IL22, IL22RA, IL20RA, IL20RB, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3, and OX40.

[0429] In a twenty-seventh aspect, the present invention provides a nucleic acid comprising a nucleotide sequence encoding an IL10RB protein modified as described above. Preferably, the nucleic acid further comprises the nucleotide sequence shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9, or a variant thereof. The variant comprises one or more of the following: a C>T mutation at position 33276503, a G>A mutation at position 33276818, or a TTTTGGG>TTG mutation at positions 33277048-33277053.

[0430] Preferably, the mRNA transcribed from the nucleic acid comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 61; B) the identity with the nucleotide sequence shown in SEQ ID NO: 61 is at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99%; C) the difference from the nucleotide sequence shown in SEQ ID NO: 61 is no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 61, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0431] In a twenty-eighth aspect, the present invention provides a polypeptide encoded by the genetically modified endogenous IL10RB gene or the aforementioned nucleic acid.

[0432] In a twenty-ninth aspect, the present invention provides a humanized IL10RB protein, said humanized IL10RB protein comprising all or part of human IL10RB protein.

[0433] Preferably, the humanized IL10RB protein comprises all or part of the signal peptide, extracellular domain, transmembrane domain and / or cytoplasmic domain of the human IL10RB protein.

[0434] In one specific embodiment of the present invention, the humanized IL10RB protein comprises all or part of the extracellular domain of human IL10RB protein, preferably comprising at least 50 consecutive amino acids of the extracellular domain of human IL10RB protein, for example, comprising at least 50, 70, 90, 100, 150, 170, 190, 198, 200, or 201 consecutive amino acids.

[0435] Preferably, the humanized IL10RB protein comprises the amino acid sequence shown in positions 21-218 of SEQ ID NO: 57; or, comprises an amino acid sequence with an identity of at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown in positions 21-218 of SEQ ID NO: 57; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 21-218 of SEQ ID NO: 57 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with an identity of positions 21-218 of SEQ ID NO: 57, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0436] Preferably, the humanized IL10RB protein comprises all or part of the amino acid sequence encoded by exons 1 to 7 of the human IL10RB gene. More preferably, it comprises all or part of the amino acid sequence encoded by any one, two, three or more, or a combination of two or more consecutive exons 1 to 7. More preferably, it comprises all or part of the amino acid sequence encoded by exons 2 to 6. More preferably, the amino acid sequences encoding a portion of exon 2, all of exons 3 to 5, and a portion of exon 6 are included. More preferably, the amino acid sequences encoding a portion of exons 2 to 6 are included. The exon 2 portion comprises at least 50 bp of nucleotide sequence, for example, at least 50, 70, 90, 100, 110, 111, 112, 113, 114, 115, 120, or 124 bp. Preferably, the exon 2 portion comprises a nucleotide sequence from the first 5 (e.g., 1, 2, 3, 4, 5) amino acids encoding the N-terminus of the extracellular domain to the last nucleotide of exon 2, and includes a nucleotide sequence encoding at least 5 consecutive amino acids of the extracellular domain of the human IL10RB protein. The nucleotide sequence contains at least 5, 7, 9, 10, 15, 20, 25, 30, 31, 32, 33, 34, 35, 38, and 39 consecutive amino acids, and the exon 6 portion contains at least 2 bp of nucleotide sequence, for example, at least 2, 3, 4, 5, 6, 7, 8, 10, 20, 50, 70, 90, 100, 120, 150, and 158 bp of nucleotide sequence; preferably, the exon 6 portion contains a nucleotide sequence from the first nucleotide of exon 6 to the C-terminus encoding the extracellular domain of 1-5 amino acids (e.g., 1, 2, 3, 4, and 5), and contains a nucleotide sequence encoding the extracellular domain of human IL10RB protein of at least 1 consecutive amino acid, for example, a nucleotide sequence containing at least 1, 2, 3, 4, and 5 consecutive amino acids.

[0437] Preferably, the humanized IL10RB protein further comprises all or part of the non-human animal IL10RB protein, and more preferably all or part of the signal peptide, extracellular domain, transmembrane domain and / or cytoplasmic domain of the non-human animal IL10RB protein.

[0438] In one specific embodiment of the present invention, the humanized IL10RB protein further comprises the amino acid sequence shown in positions 1-22 and 221-351 of SEQ ID NO: 56; or, comprises an amino acid sequence with an identity of at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown in positions 1-22 and 221-351 of SEQ ID NO: 56; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 1-22 and 221-351 of SEQ ID NO: 56 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with an identity of positions 1-22 and 221-351 of SEQ ID NO: 56, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0439] In one specific embodiment of the present invention, the amino acid sequence of the humanized IL10RB protein comprises any one of the following groups: A) the amino acid sequence shown in SEQ ID NO: 62; B) the amino acid sequence being at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identical to the amino acid sequence shown in SEQ ID NO: 62; C) the amino acid sequence differing from the amino acid sequence shown in SEQ ID NO: 62 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or D) the amino acid sequence shown in SEQ ID NO: 62, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0440] Preferably, the non-human animal can be selected from rodents, pigs, rabbits, monkeys, or any other non-human animal that can be genetically edited to produce humanized genes. Further limitations on the type of non-human animal are the same as in the sixth aspect of this invention.

[0441] In a thirtieth aspect of the present invention, a humanized IL10RB gene is provided, wherein the humanized IL10RB gene comprises a portion of the human IL10RB gene.

[0442] Preferably, the humanized IL10RB gene comprises all or part of exons 1 to 7 of the human IL10RB gene. More preferably, the humanized IL10RB gene comprises all or part of any one, two, three or more, or consecutive combinations of two or more exons 1 to 7 of the human IL10RB gene. More preferably, it comprises all or part of exons 2 to 6. Even more preferably, the humanized IL10RB gene comprises a portion of exon 2, all of exons 3 to 5, and a portion of exon 6 of the human IL10RB gene, preferably also comprising intron 2 and / or intron 5, and more preferably comprising any intron between exons 2 and 6. In one specific embodiment of the present invention, the humanized IL10RB gene comprises a portion of exons 2 to 6 of the human IL10RB gene, wherein the portion of exon 2 comprises at least 50 bp of nucleotide sequence, for example, at least 50, 70, 90, 100, 110, 111, 112, 113, 114, 115, 120, or 124 bp of nucleotide sequence; preferably, the portion of exon 2 comprises a nucleotide sequence from the first to fifth (e.g., 1, 2, 3, 4, 5) amino acids encoding the N-terminus of the extracellular domain to the last nucleotide of exon 2, comprising a nucleotide sequence encoding at least five consecutive amino acids of the extracellular domain of the human IL10RB protein, for example, at least 5, The nucleotide sequence comprises 7, 9, 10, 15, 20, 25, 30, 31, 32, 33, 34, 35, 38, and 39 consecutive amino acids, and the exon 6 portion comprises at least 5 bp of nucleotide sequence, for example, at least 5, 6, 7, 8, 10, 20, 50, 70, 90, 100, 120, 150, and 158 bp of nucleotide sequence; preferably, the exon 6 portion comprises a nucleotide sequence from the first nucleotide of exon 6 to the C-terminus encoding the extracellular domain of 1-5 amino acids (e.g., 1, 2, 3, 4, and 5), and a nucleotide sequence encoding the extracellular domain of human IL10RB protein of at least 1 consecutive amino acid, for example, a nucleotide sequence comprising at least 1, 2, 3, 4, and 5 consecutive amino acids.

[0443] In one specific embodiment of the present invention, the humanized IL10RB gene comprises the nucleotide sequence or a variant thereof shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9; or, comprises a nucleotide sequence or a variant thereof with an identity of at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 95%, 90%, 91%, 92%, 93%, 94%, 95%, 95%, 90%, 95%, 90%, 90%, 91%, 92%, 93%, 94%, 95%, 95%, 90 ... 6%, 97%, 98%, or at least 99%; or, containing a nucleotide sequence differing from the nucleotide sequence shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or its variants by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or, containing a nucleotide sequence having the nucleotide sequence shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or its variants, including substitutions, deletions, and / or insertions of one or more nucleotides. The variants include one or more of the following: C>T mutation at position 33276503, G>A mutation at position 33276818, or TTTTGGG>TTG mutation at positions 33277048-33277053.

[0444] Preferably, the humanized IL10RB gene contains all or part of the nucleotide sequence encoding the human IL10RB protein. Further preferably, the nucleotide sequence comprises all or part of the extracellular domain encoding the human IL10RB protein, and more preferably, the nucleotide sequence encoding positions 21-218 of SEQ ID NO: 57; or, the nucleotide sequence having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence encoding positions 21-218 of SEQ ID NO: 57; or, the nucleotide sequence differing from the amino acid sequence encoding positions 21-218 of SEQ ID NO: 57 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, the nucleotide sequence having the nucleotide sequence encoding positions 21-218 of SEQ ID NO: 57, including nucleotide sequences with substitutions, deletions, and / or insertions of one or more nucleotides.

[0445] Preferably, the humanized IL10RB gene further comprises all or part of the non-human animal IL10RB gene. More preferably, it comprises all or part of exons 1 to 7 of the non-human animal IL10RB gene, and more preferably, it comprises all of exon 1, part of exon 2, part of exon 6, and / or all of exon 7 of the non-human animal IL10RB gene.

[0446] In one specific embodiment of the present invention, the humanized IL10RB gene further comprises a nucleotide sequence encoding positions 1-22 and 221-351 of SEQ ID NO: 56; or, comprises a nucleotide sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity to the nucleotide sequence encoding positions 1-22 and 221-351 of SEQ ID NO: 56; or, comprises a nucleotide sequence differing from the amino acid sequence encoding positions 1-22 and 221-351 of SEQ ID NO: 56 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises a nucleotide sequence having a nucleotide sequence encoding positions 1-22 and 221-351 of SEQ ID NO: 56, including nucleotide sequences with substitutions, deletions, and / or insertions of one or more nucleotides.

[0447] In one specific embodiment of the present invention, the nucleotide sequence of the humanized IL10RB gene comprises any one of the following groups: A) the transcribed mRNA is the nucleotide sequence shown in SEQ ID NO: 61; B) the transcribed mRNA is at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identical to the nucleotide sequence shown in SEQ ID NO: 61; C) the transcribed mRNA differs from the nucleotide sequence shown in SEQ ID NO: 61 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) the transcribed mRNA has the nucleotide sequence shown in SEQ ID NO: 61, including nucleotide sequences with substitutions, deletions, and / or insertions of one or more nucleotides.

[0448] Preferably, the humanized IL10RB gene encodes the aforementioned humanized IL10RB protein.

[0449] Preferably, the humanized IL10RB gene further includes a specific inducer or repressor. More preferably, the specific inducer or repressor can be a conventionally induced or repressible substance. In one specific embodiment of the present invention, the specific inducer is selected from the tetracycline system (Tet-Off System / Tet-On System) or the tamoxifen system.

[0450] Preferably, the non-human animal can be selected from rodents, pigs, rabbits, monkeys, or any other non-human animal that can be genetically edited to produce humanized genes. Further limitations on the type of non-human animal are the same as in the sixth aspect of this invention.

[0451] In a thirty-first aspect, the present invention provides a targeting vector comprising a donor nucleotide sequence comprising one of the following: A) a nucleotide sequence encoding a humanized IL10RB protein; B) a nucleotide sequence encoding a human IL10RB protein, preferably comprising all or part of a nucleotide sequence encoding a signal peptide, extracellular domain, transmembrane domain, and / or cytoplasmic domain of the human IL10RB protein, more preferably comprising all or part of a nucleotide sequence encoding the extracellular domain of the human IL10RB protein, more preferably comprising a nucleotide sequence encoding at least 50 consecutive amino acids of the extracellular domain of the human IL10RB protein, and even more preferably comprising a nucleotide sequence encoding the amino acid sequence shown at positions 21-218 of SEQ ID NO: 57; C) a nucleotide sequence of a humanized IL10RB gene; or, D) a portion of the human IL10RB gene, preferably comprising all or part of exons 1 to 7 of the human IL10RB gene. More preferably, it comprises all or part of any one, two, three or more, or a combination of two or more consecutive exons 1 to 7. More preferably, it contains all or part of exons 2 to 6 of the human IL10RB gene; even more preferably, it contains a portion of exon 2, all of exons 3 to 5, and a portion of exon 6, particularly a portion of exons 2 to 6. The exon 2 portion contains at least 50 bp of nucleotide sequence, for example, at least 50, 70, 90, 100, 110, 111, 112, 113, 114, 115, 120, or 124 bp. Preferably, the exon 2 portion contains a nucleotide sequence from the first 5 amino acids (e.g., 1, 2, 3, 4, 5) encoding the N-terminus of the extracellular domain to the last nucleotide of exon 2, containing at least 5 consecutive amino acids encoding the extracellular domain of the human IL10RB protein. The nucleotide sequence, for example, contains at least 5, 7, 9, 10, 15, 20, 25, 30, 31, 32, 33, 34, 35, 38, or 39 consecutive amino acids, and the exon 6 portion contains at least 5 bp of nucleotide sequence, for example, contains at least 5, 6, 7, 8, 10, 20, 50, 70, 90, 100, 120, 150, or 158 bp of nucleotide sequence; preferably, the exon 6 portion contains a nucleotide sequence from the first nucleotide of exon 6 to the C-terminus encoding the extracellular domain of 1-5 (e.g., 1, 2, 3, 4, 5) amino acids, and contains a nucleotide sequence encoding at least one consecutive amino acid of the extracellular domain of human IL10RB protein, for example, a nucleotide sequence containing at least 1, 2, 3, 4, or 5 consecutive amino acids.Further preferably, the donor nucleotide sequence comprises the nucleotide sequence shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or a variant thereof; or, comprises a nucleotide sequence with an identity of at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the nucleotide sequence shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or a variant thereof; or, comprises a nucleotide sequence with an identity of at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the nucleotide sequence shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or a variant thereof. The nucleotide sequence shown at positions 68405-33288111 or its variants differ by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2 or no more than 1 nucleotide; or, it contains a nucleotide sequence shown at positions 33268405-33288111 or its variants with NCBI gene accession number NC_000021.9, including substitutions, deletions and / or insertions of one or more nucleotides, wherein the variants include one or more of the following: C>T mutation at position 33276503, G>A mutation at position 33276818, or TTTTGGG>TTG mutation at positions 33277048-33277053.

[0452] Preferably, the humanized IL10RB protein is the aforementioned humanized IL10RB protein.

[0453] Preferably, the humanized IL10RB gene is the aforementioned humanized IL10RB gene.

[0454] Preferably, the targeting vector further comprises a 5' arm (5' homologous arm) and / or a 3' arm (3' homologous arm).

[0455] Preferably, the 5' arm is a DNA fragment homologous to the 5' end of the conversion region to be modified, selected from 100-10000 nucleotides in length from the genomic DNA of the non-human animal IL10RB gene. Preferably, the 5' arm has at least 90% homology with the nucleotide in NCBI accession number NC_000082.7. More preferably, the 5' arm sequence comprises SEQ ID NO: 58.

[0456] Preferably, the 3' arm is a second DNA fragment homologous to the 3' end of the conversion region to be modified, and is selected from 100-10000 nucleotides in length from the genomic DNA of the non-human animal IL10RB gene; preferably, the 3' arm has at least 90% homology with the nucleotide in NCBI accession number NC_000082.7; more preferably, the 3' arm sequence contains SEQ ID NO: 59.

[0457] Preferably, the targeting vector further comprises SEQ ID NO: 63, 64, 65 and / or 66.

[0458] Preferably, the conversion region to be altered is located on exons 2 to 6 of the non-human animal IL10RB gene.

[0459] Preferably, the targeting vector further comprises a marker gene, a resistance gene for positive clone selection, and / or a specific recombination system. The relevant limitations regarding the marker gene, the resistance gene for positive clone selection, and / or the specific recombination system are the same as in aspect eight of the present invention.

[0460] In a thirty-second aspect, the present invention provides a cell comprising the targeting vector described in the thirty-first aspect.

[0461] In a thirty-third aspect, the present invention provides the application of the targeting vector described in the thirty-first aspect and / or the cells described in the thirty-second aspect in IL10RB gene editing, preferably, the application including but not limited to knockout, insertion or replacement.

[0462] In a thirty-fourth aspect, the present invention provides a non-human animal with humanized IL10RB gene expression, wherein the non-human animal expresses human or humanized IL10RB protein.

[0463] Preferably, the humanized IL10RB protein described above is expressed in the non-human animal.

[0464] Preferably, the expression of endogenous IL10RB protein in the non-human animal is reduced or absent.

[0465] Preferably, the non-human animal genome contains a portion of the human IL10RB gene or a humanized IL10RB gene, more preferably it contains the aforementioned humanized IL10RB gene.

[0466] Preferably, a portion of the human IL10RB gene or the nucleotide sequence of the humanized IL10RB gene is operatively linked to an endogenous regulatory element at an endogenous IL10RB gene locus in at least one chromosome.

[0467] Preferably, the non-human animal is constructed by introducing a donor nucleotide sequence into the non-human animal IL10RB gene locus. Preferably, the limitation of the donor nucleotide sequence is the same as in aspect thirty-first of the present invention.

[0468] Preferably, the non-human animal is constructed using the aforementioned targeting vector.

[0469] According to some embodiments of the present invention, the non-human animal further comprises other gene modifications, said other genes being selected from at least one of IL22, IL22RA1, IL20RA, IL20RB, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3, and OX40.

[0470] Preferably, the IL22 gene is a humanized IL22 gene; more preferably, the humanized IL22 gene is any of the humanized IL22 genes described in this invention.

[0471] Preferably, the IL22RA1 gene is a humanized IL22RA1 gene; more preferably, the humanized IL22RA1 gene is any of the humanized IL22RA1 genes described in this invention.

[0472] Preferably, the IL20RA gene is a humanized IL20RA gene; more preferably, the humanized IL20RA gene is any of the humanized IL20RA genes described in this invention.

[0473] Preferably, the IL20RB gene is a humanized IL20RB gene; more preferably, the humanized IL20RB gene is any of the humanized IL20RB genes described in this invention.

[0474] According to some embodiments of the present invention, the partial or humanized IL10RB gene of the human IL10RB gene and / or the other genes are homozygous or heterozygous for the endogenously replaced locus.

[0475] Preferably, the humanized IL10RB gene further includes a specific inducer or repressor; more preferably, the specific inducer or repressor can be a conventionally induced or repressible substance. In one specific embodiment of the present invention, the specific inducer is selected from the tetracycline system (Tet-Off System / Tet-On System) or the tamoxifen system.

[0476] Preferably, the non-human animal can be selected from rodents, pigs, rabbits, monkeys, or any other non-human animal that can be genetically edited to produce humanized genes. Further limitations on the type of non-human animal are the same as in the sixth aspect of this invention.

[0477] In a thirty-fifth aspect of the present invention, a method for constructing a non-human animal with humanized IL10RB gene is provided, wherein the non-human animal expresses human or humanized IL10RB protein.

[0478] Preferably, the humanized IL10RB protein described above is expressed in the non-human animal.

[0479] Preferably, the genome of at least one cell of the non-human animal contains a portion of the human IL10RB gene or a humanized IL10RB gene, more preferably the aforementioned humanized IL10RB gene.

[0480] Preferably, the non-human animal is a non-human animal with the IL10RB gene humanized as described above.

[0481] Preferably, the construction method includes introducing a donor nucleotide sequence into the IL10RB gene locus in a non-human animal. Preferably, the limitation of the donor nucleotide sequence is the same as in aspect thirty-first of the present invention.

[0482] Preferably, a portion or humanized IL10RB gene of the human IL10RB gene is operatively linked to an endogenous IL10RB regulatory element on at least one chromosome.

[0483] Preferably, the partial or humanized IL10RB gene of the human IL10RB gene is regulated in non-human animals by a regulatory element. More preferably, the regulatory element can be endogenous or exogenous.

[0484] Preferably, the control elements include, but are not limited to, a promoter, a 5'UTR, and / or a 3'UTR.

[0485] In one specific embodiment of the present invention, the endogenous regulatory element is derived from the non-human animal IL10RB gene, and the exogenous regulatory element is derived from the human IL10RB gene.

[0486] Preferably, the insertion site is located after the endogenous regulatory element of the IL10RB gene.

[0487] Preferably, the introduction is a substitution, insertion, or transgenesis. Optionally, the introduction of the non-human animal IL10RB gene locus is a substitution of the corresponding region in the non-human animal. More preferably, all or part of exons 2 to 6 of the non-human animal IL10RB gene are substituted. More preferably, part of exon 2, all of exon 3, all of exon 4, all of exon 5, and part of exon 6 of the non-human animal IL10RB gene are substituted. Even more preferably, part of exons 2 to 6 of the non-human animal IL10RB gene are substituted.

[0488] Preferably, the nucleotide sequence encoding the amino acids at positions 23-220 of SEQ ID NO:56 is replaced.

[0489] Preferably, the non-human animal is constructed using gene editing technology to humanize the IL10RB gene. The gene editing technology includes gene targeting technology using embryonic stem cells, CRISPR / Cas9 technology, zinc finger nuclease technology, transcription activator-like effector nuclease technology, homing endonuclease or other molecular biology techniques.

[0490] In one specific embodiment of the present invention, the construction method includes modifying the coding frame of the non-human animal IL10RB gene by inserting a nucleotide sequence encoding the human or humanized IL10RB protein, or a portion of the human IL10RB gene, or a nucleotide sequence of the humanized IL10RB gene, after the endogenous regulatory element of the non-human animal IL10RB gene. The modified coding frame of the non-human animal IL10RB gene can be a knockout of the functional region of the non-human animal IL10RB gene or the insertion of a sequence, causing the non-human animal IL10RB protein to be unexpressed, underexpressed, or non-functional. More preferably, the modified coding frame of the non-human animal IL10RB gene can be a knockout of all or part of exons 2 to 6 of the non-human animal IL10RB gene, preferably a portion of exon 2, all of exon 3, all of exon 4, all of exon 5, and a portion of exon 6, and even more preferably a portion of exons 2 to 6.

[0491] Preferably, the above-mentioned targeting vector is used to construct non-human animals.

[0492] In one specific embodiment of the present invention, the construction method includes introducing the above-mentioned targeting vector into non-human animal cells (preferably embryonic stem cells), screening out the correct positive clone cells and introducing them into the isolated blastocysts, culturing the blastocysts, and then transplanting the cultured blastocysts into the oviducts of female non-human animals, allowing them to develop, and identifying and screening to obtain non-human animals with humanized IL10RB genes.

[0493] According to some embodiments of the present invention, the construction method further includes: mating non-human animals with humanized IL10RB genes with other genetically modified non-human animals, in vitro fertilization or direct gene editing, and screening to obtain multi-gene-modified non-human animals.

[0494] Preferably, the other genes are non-human animals modified with at least one of the following genes: IL22, IL22RA1, IL20RA, IL20RB, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3, and OX40.

[0495] Preferably, the non-human animal also expresses at least one of the following human or humanized proteins: IL22, IL22RA1, IL20RA, IL20RB, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3, and OX40.

[0496] Preferably, the IL22RA1 gene is a humanized IL22RA1 gene; more preferably, the humanized IL22RA1 gene is any of the humanized IL22RA1 genes described in this invention.

[0497] Preferably, the IL22 gene is a humanized IL22 gene; more preferably, the humanized IL22 gene is any of the humanized IL22 genes described in this invention.

[0498] Preferably, the IL20RA gene is a humanized IL20RA gene; more preferably, the humanized IL20RA gene is any of the humanized IL20RA genes described in this invention.

[0499] Preferably, the IL20RB gene is a humanized IL20RB gene; more preferably, the humanized IL20RB gene is any of the humanized IL20RB genes described in this invention.

[0500] Preferably, the IL22RA1 protein is a humanized IL22RA1 protein; more preferably, the humanized IL22RA1 protein is any of the humanized IL22RA1 proteins described in this invention.

[0501] Preferably, the IL22 protein is a humanized IL22 protein; more preferably, the humanized IL22 protein is any of the humanized IL22 proteins described in this invention.

[0502] Preferably, the IL20RA protein is a humanized IL20RA protein; more preferably, the humanized IL20RA protein is any of the humanized IL20RA proteins described in this invention.

[0503] Preferably, the IL20RB protein is a humanized IL20RB protein; more preferably, the humanized IL20RB protein is any of the humanized IL20RB proteins described in this invention.

[0504] Preferably, in the genome of the multi-gene modified non-human animal, each of the modified genes is homozygous or heterozygous for the endogenous replaced locus.

[0505] Preferably, the non-human animal can be selected from rodents, pigs, rabbits, monkeys, or any other non-human animal that can be genetically edited to produce humanized genes. Further limitations on the type of non-human animal are the same as in the sixth aspect of this invention.

[0506] In a thirty-sixth aspect of the present invention, a non-human animal genome is provided, wherein the non-human animal or the non-human animal genome comprises a genetically modified endogenous IL20RA gene encoding a modified IL20RA protein, and the non-human animal expresses the modified IL20RA protein.

[0507] Preferably, the non-human animal or non-human animal genome contains a nucleotide sequence that replaces the corresponding region encoding the non-human animal endogenous IL20RA with a nucleotide sequence encoding the human IL20RA region.

[0508] Preferably, the modified IL20RA protein contains an extracellular domain that is substantially the same as that of the human IL20RA protein.

[0509] Preferably, all or part of the extracellular domain comprises the amino acid sequence shown in positions 30-250, 34-250, 30-247, or 34-247 of SEQ ID NO: 83; or, comprises an amino acid sequence with an identity of at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with ...10, 9, 8, 7, 6, 5, 4, 3, 2, or at least 1 amino acid with the amino acid sequence shown in positions 30-250, 34-250, 30-247, or 34-247 of SEQ ID NO: 83; or, comprises an amino acid sequence with an identity of at least 10, 9, 8, 7, 6, 5, 4, 3, 2, or at least 1 amino acid with the amino acid sequence shown in positions 30-250, 34-250, 30-247, or 34-247 of SEQ ID NO: 83; or, comprises an amino acid sequence with an identity of at least 10, 9, 8, 7, NO:83 indicates an amino acid sequence including substitutions, deletions, and / or insertions of one or more amino acid residues, as shown in positions 30-250, 34-250, 30-247, or 34-247.

[0510] Preferably, the extracellular domain comprises 0-10 amino acid residues (e.g., 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10) at the N-terminus and / or C-terminus of the extracellular domain of the non-human endogenous IL20RA protein, more preferably comprising the amino acid sequence shown at positions 33-36 of SEQ ID NO:82; or, comprising amino acid sequences with at least 77%, 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown at positions 33-36 of SEQ ID NO:82; or, comprising amino acid sequences with differences not exceeding 10, 9, 8, 7, 6, 5, 4, 3, 2, or not exceeding 1 amino acid from .... The amino acid sequence shown at positions 33-36 of NO:82 includes substitutions, deletions, and / or insertions of one or more amino acid residues.

[0511] Preferably, the non-human animal is a rodent, preferably a rat or mouse.

[0512] Preferably, the modified IL20RA protein contains a cytoplasmic domain that is substantially the same as that of the non-human animal endogenous IL20RA protein.

[0513] Preferably, the amino acid sequence of the cytoplasmic domain comprises the amino acid sequence shown in positions 275-546 of SEQ ID NO: 82; or, comprises an amino acid sequence with an identity of at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown in positions 275-546 of SEQ ID NO: 82; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 275-546 of SEQ ID NO: 82 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with an identity of positions 275-546 of SEQ ID NO: 82, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0514] Preferably, the modified IL20RA protein contains a transmembrane domain that is substantially the same as that of the non-human endogenous IL20RA protein.

[0515] Preferably, the amino acid sequence of the transmembrane domain comprises the amino acid sequence shown in positions 254-274 of SEQ ID NO: 82; or, comprises an amino acid sequence with an identity of at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown in positions 254-274 of SEQ ID NO: 82; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 254-274 of SEQ ID NO: 82 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with an identity of positions 254-274 of SEQ ID NO: 82, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0516] Preferably, the modified IL20RA protein comprises a non-human animal endogenous cytoplasmic domain, a non-human animal endogenous transmembrane domain, and a human extracellular domain.

[0517] Preferably, the modified IL20RA protein has an amino acid sequence comprising the amino acid sequence shown in SEQ ID NO: 91; or, having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in SEQ ID NO: 91; or, differing from the amino acid sequence shown in SEQ ID NO: 91 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, having an amino acid sequence similar to that shown in SEQ ID NO: 91, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0518] Preferably, the replaced nucleotide sequence encoding the corresponding region of non-human endogenous IL20RA comprises all or part of the nucleotide sequence encoding the extracellular domain of the non-human endogenous IL20RA protein, more preferably comprising the nucleotide sequence encoding positions 37-253 or 33-253 of SEQ ID NO: 82; or, comprising a nucleotide sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence encoding positions 37-253 or 33-253 of SEQ ID NO: 82; or, comprising a nucleotide sequence with a difference of no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide from ... NO:82 The nucleotide sequence shown at positions 37-253 or 33-253 includes a nucleotide sequence with substitutions, deletions and / or insertions of one or more nucleotides, more preferably a partial nucleotide sequence containing exons 2 to 6 of a non-human animal endogenous IL20RA sequence.

[0519] Preferably, the nucleotide sequence encoding the human IL20RA region comprises a portion of the human IL20RA gene; preferably, it comprises a portion of exon 2, all of exon 3, all of exon 4, all of exon 5, and a portion of exon 6 of the human IL20RA gene; more preferably, it comprises a portion of exons 2 to 6 of the human IL20RA gene, wherein the portion of exon 2 comprises at least 20 bp of continuous nucleotide sequence, such as at least 20, 30, 40, 50, 60, 70, 80, 90, 100, 110, 120, 125, 130, or 136 bp of continuous nucleotide sequence, preferably comprising all or part of the nucleotide sequence encoding the extracellular domain in exon 2. The portion of exon 6 contains at least 5 bp of continuous nucleotide sequences, such as 5, 10, 15, 20, 21, 22, 23, 24, 25, 26, 30, 40, 50, 60, 70, 80, 90, 100, 110, 120, 130 or 140 bp of continuous nucleotide sequences, preferably containing all or part of the nucleotide sequence encoding the extracellular domain in exon 6.

[0520] Preferably, the nucleotide sequence encoding the human IL20RA region comprises the nucleotide sequence shown in positions 137004735-137017092 of NCBI Gene Accession Number NC_000006.12; or, it comprises a nucleotide sequence with an identity of at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99. 8% or at least 99% of the nucleotide sequence; or, a nucleotide sequence that differs from the nucleotide sequence shown in positions 137004735-137017092 of NCBI Gene Accession Number NC_000006.12 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2 or no more than 1 nucleotide; or, a nucleotide sequence that includes substitutions, deletions and / or insertions of one or more nucleotides as shown in the nucleotide sequence shown in positions 137004735-137017092 of NCBI Gene Accession Number NC_000006.12.

[0521] Preferably, the mRNA transcribed from the nucleotide sequence encoding the modified IL20RA protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 90; B) the nucleotide sequence having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 90; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 90 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 90, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0522] Preferably, the non-human animal genome includes replacing all or part of the nucleotide sequence encoding the extracellular domain of the endogenous IL20RA protein in the non-human animal genome with a nucleotide sequence encoding an extracellular domain substantially identical to that of the human IL20RA protein.

[0523] Preferably, the non-human animal genome includes replacing all or part of the nucleotide sequence encoding the extracellular domain of the endogenous IL20RA protein in the non-human animal genome with a nucleotide sequence encoding the human IL20RA region.

[0524] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding positions 37-253 or 33-253 of SEQ ID NO: 82 in the non-human animal genome with a nucleotide sequence encoding the human IL20RA region (e.g., genomic DNA sequence, cDNA sequence or CDS sequence).

[0525] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding positions 37-253 or 33-253 of SEQ ID NO: 82 in the non-human animal genome with a portion of exon 2 to exon 6 of the human IL20RA gene.

[0526] Preferably, the non-human animal genome includes replacing parts of exons 2 to 6 of the non-human animal endogenous IL20RA gene in the non-human animal genome with parts of exons 2 to 6 of the human IL20RA gene.

[0527] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding positions 37-253 of SEQ ID NO: 82 in the non-human animal genome with the nucleotide sequence encoding positions 34-250 of SEQ ID NO: 83.

[0528] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding positions 33-253 of SEQ ID NO: 82 in the non-human animal genome with the nucleotide sequence encoding positions 30-250 of SEQ ID NO: 83.

[0529] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding SEQ ID NO: 82 in the non-human animal genome with the nucleotide sequence encoding SEQ ID NO: 91.

[0530] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding positions 37-253 of SEQ ID NO: 82 in the non-human animal genome with the nucleotide sequence shown in positions 137004735-137017092 of NCBI accession number NC_000006.12.

[0531] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding SEQ ID NO: 82 in the non-human animal genome with the nucleotide sequence shown in SEQ ID NO: 90.

[0532] Preferably, the expression of the modified IL20RA protein is controlled by endogenous IL20RA regulatory elements (e.g., promoter, 5'UTR and / or 3'UTR).

[0533] Preferably, the non-human animal genome is homozygous or heterozygous for the genetically modified endogenous IL20RA gene.

[0534] Preferably, the non-human animal genome contains endogenous genes encoding human or modified or wild-type other genes, and preferably, the other genes are selected from at least one of IL22, IL22RA1, IL10RB, IL20RB, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3 and OX40.

[0535] Preferably, the non-human animal genome contains an endogenous IL10RB locus encoding a human, modified, or wild-type IL10RB protein.

[0536] In one specific embodiment of the present invention, the non-human animal genome contains a genetically modified endogenous IL10RB locus encoding a modified IL10RB protein, and the non-human animal expresses the modified IL10RB protein.

[0537] Preferably, in the genetically modified endogenous IL10RB locus, the nucleotide sequence encoding the corresponding segment of non-human endogenous IL10RB is replaced with the nucleotide sequence encoding the human IL10RB segment.

[0538] Preferably, the modified IL10RB protein comprises the amino acid sequence shown in SEQ ID NO: 62; or, comprises an amino acid sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in SEQ ID NO: 62; or, comprises an amino acid sequence differing from the amino acid sequence shown in SEQ ID NO: 62 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence similar to that shown in SEQ ID NO: 62, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0539] Preferably, the nucleotide sequence encoding the human IL10RB region includes a portion of the human IL10RB gene; more preferably, it includes a portion of exon 2, all of exon 3, all of exon 4, all of exon 5, and a portion of exon 6 of the human IL10RB gene; even more preferably, it includes portions of exons 2 to 6.

[0540] Preferably, the nucleotide sequence encoding the human IL10RB region comprises the nucleotide sequence shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or a variant thereof; or, comprises a nucleotide sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity to the nucleotide sequence shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or a variant thereof; or, comprises a nucleotide sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity to the nucleotide sequence shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or a variant thereof. The nucleotide sequence shown at positions 33268405-33288111 or its variants differ by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2 or no more than 1 nucleotide; or, a nucleotide sequence containing the nucleotide sequence shown at positions 33268405-33288111 or its variants with NCBI gene accession number NC_000021.9, including substitutions, deletions and / or insertions of one or more nucleotides, wherein the variants include one or more of the following: C>T mutation at position 33276503, G>A mutation at position 33276818, or TTTTGGG>TTG mutation at positions 33277048-33277053;

[0541] Preferably, the mRNA transcribed from the nucleotide sequence encoding the modified IL10RB protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 61; B) the nucleotide sequence having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 61; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 61 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 61, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0542] Preferably, the non-human animal genome contains an endogenous IL20RB locus encoding a human, modified, or wild-type IL20RB protein.

[0543] In one specific embodiment of the present invention, the non-human animal genome contains a genetically modified endogenous IL20RB locus encoding a modified IL20RB protein, and the non-human animal expresses the modified IL20RB protein.

[0544] Preferably, in the genetically modified endogenous IL20RB locus, the nucleotide sequence encoding the corresponding segment of non-human animal endogenous IL20RB is replaced with the nucleotide sequence encoding the human IL20RB segment.

[0545] Preferably, the modified IL20RB protein has an amino acid sequence comprising the amino acid sequence shown in SEQ ID NO: 121; or, having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in SEQ ID NO: 121; or, differing from the amino acid sequence shown in SEQ ID NO: 121 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, having an amino acid sequence similar to that shown in SEQ ID NO: 121, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0546] Preferably, the nucleotide sequence encoding the human IL20RB region includes a portion of the human IL20RB gene; more preferably, it includes a portion of exon 2, all of exon 3, all of exon 4, all of exon 5, and a portion of exon 6 of the human IL20RB gene; even more preferably, it includes portions of exons 2 to 6.

[0547] Preferably, the nucleotide sequence encoding the human IL20RB region comprises the nucleotide sequence shown in positions 136980507-136995463 of NCBI Gene Accession Number NC_000003.12; or, it comprises a nucleotide sequence with an identity of at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 9 ... 8% or at least 99% of the nucleotide sequence; or, a nucleotide sequence that differs from the nucleotide sequence shown in positions 136980507-136995463 of NCBI Gene Accession Number NC_000003.12 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2 or no more than 1 nucleotide; or, a nucleotide sequence that includes substitutions, deletions and / or insertions of one or more nucleotides as shown in the nucleotide sequence shown in positions 136980507-136995463 of NCBI Gene Accession Number NC_000003.12.

[0548] Preferably, the mRNA transcribed from the nucleotide sequence encoding the modified IL20RB protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 120; B) the nucleotide sequence having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 120; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 120 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 120, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0549] Preferably, the non-human animal genome contains an endogenous IL10 locus encoding a human, modified, or wild-type IL10 protein.

[0550] Preferably, the non-human animal or non-human animal genome contains a genetically modified endogenous IL10 locus encoding a human or modified IL10 protein, and the non-human animal expresses the human or modified IL10 protein.

[0551] Preferably, in the genetically modified endogenous IL10 locus, the nucleotide sequence of the corresponding segment of the non-human animal endogenous IL10 is replaced with the nucleotide sequence encoding the human IL10 segment.

[0552] Preferably, the nucleotide sequence encoding the human IL10 region comprises a portion of exon 1, all of exon 2, all of exon 3, all of exon 4, and a portion of exon 5 of the human IL10 gene; more preferably, it comprises portions of exons 1 to 5. The portion of exon 1 comprises at least 50 bp, for example, at least 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, 160, 165, 170, 180, 190, 200, 220, or 224 bp of continuous nucleotide sequence. The portion of exon 5 contains at least 50 bp, such as at least 50, 60, 70, 80, 90, 93, 100, 200, 300, 400, 500, 600, 700, 800, 900, 1000, 1100, 1120, or 1127 bp of continuous nucleotide sequence.

[0553] Preferably, the non-human animal genome contains an endogenous IL10RA locus encoding a human, modified, or wild-type IL10RA protein.

[0554] Preferably, the non-human animal or the non-human animal genome contains a genetically modified endogenous IL10RA locus encoding a modified IL10RA protein, and the non-human animal expresses the modified IL10RA protein.

[0555] Preferably, in the genetically modified endogenous IL10RA locus, the nucleotide sequence encoding the corresponding segment of non-human animal endogenous IL10RA is replaced with the nucleotide sequence encoding the human IL10RA segment.

[0556] Preferably, the modified IL10RA protein comprises a cytoplasmic domain, a transmembrane domain, and an extracellular domain.

[0557] Preferably, the modified IL10RA protein further comprises a signal peptide.

[0558] Preferably, the IL10RA protein contains a signal peptide that is substantially the same as that of the human IL10RA protein.

[0559] Preferably, the IL10RA protein contains an extracellular domain that is substantially the same as that of the human IL10RA protein.

[0560] Preferably, the IL10RA protein contains a transmembrane domain that is substantially the same as that of the human IL10RA protein.

[0561] Preferably, the IL10RA protein contains a cytoplasmic domain that is substantially the same as that of the human IL10RA protein.

[0562] Preferably, the cytoplasmic domain also includes all or part of the cytoplasmic domain of the non-human animal endogenous IL10RA protein.

[0563] Preferably, the nucleotide sequence encoding the human IL10RA segment includes a portion of exon 1, all of exon 2, all of exon 3, all of exon 4, all of exon 5, and a portion of exon 6 of the human IL10RA gene; more preferably, it includes portions of exons 1 to 6.

[0564] Preferably, the non-human animal genome comprises any one of the following groups:

[0565] 1) Genetically modified endogenous IL20RA genes and wild-type endogenous IL20RB genes encoding modified IL20RA proteins;

[0566] 2) Genetically modified endogenous IL20RA genes, wild-type endogenous IL20RB genes, and wild-type endogenous IL19 genes encoding modified IL20RA proteins;

[0567] 3) Genetically modified endogenous IL20RA gene encoding modified IL20RA protein, genetically modified endogenous IL20RB gene encoding modified IL20RB protein, and wild-type endogenous IL19 gene;

[0568] 4) Genetically modified endogenous IL20RA gene, wild-type endogenous IL20RB gene, and wild-type endogenous IL20 gene encoding modified IL20RA protein;

[0569] 5) Genetically modified endogenous IL20RA gene encoding modified IL20RA protein, genetically modified endogenous IL20RB gene encoding modified IL20RB protein, and wild-type endogenous IL20 gene;

[0570] 6) Genetically modified endogenous IL20RA genes, wild-type endogenous IL20RB genes, and wild-type endogenous IL24 genes encoding modified IL20RA proteins;

[0571] 7) Genetically modified endogenous IL20RA gene encoding modified IL20RA protein, genetically modified endogenous IL20RB gene encoding modified IL20RB protein, and wild-type endogenous IL24 gene;

[0572] 8) Genetically modified endogenous IL20RA genes and wild-type endogenous IL10RB genes encoding modified IL20RA proteins;

[0573] 9) Genetically modified endogenous IL20RA genes, wild-type endogenous IL10RB genes, and wild-type endogenous IL26 genes encoding modified IL20RA proteins;

[0574] 10) Genetically modified endogenous IL20RA genes encoding modified IL20RA proteins, genetically modified endogenous IL10RB genes encoding human or modified IL10RB proteins, and wild-type endogenous IL26 genes.

[0575] In a thirty-seventh aspect of the present invention, a non-human animal is provided, the non-human animal comprising a genetically modified endogenous IL20RA gene encoding a modified IL20RA protein, the non-human animal expressing the modified IL20RA protein.

[0576] Preferably, the relevant limitations on the modified IL20RA protein and the genetically modified endogenous IL20RA gene are the same as those in aspect thirty-six of the present invention.

[0577] Preferably, the non-human animal comprises the non-human animal genome described in aspect thirty-six.

[0578] In a thirty-eighth aspect of the present invention, a method for preparing a non-human animal is provided, wherein the non-human animal or the genome of the non-human animal contains a genetically modified endogenous IL20RA gene encoding a modified IL20RA protein, and the non-human animal expresses the modified IL20RA protein.

[0579] Preferably, the genome of the non-human animal or non-human animal contains a nucleotide sequence that replaces the corresponding region encoding the endogenous IL20RA region of the non-human animal with a nucleotide sequence encoding the human IL20RA region.

[0580] Preferably, the modified IL20RA protein comprises a cytoplasmic domain, a transmembrane domain, and an extracellular domain.

[0581] Preferably, the modified IL20RA protein contains an extracellular domain that is substantially the same as that of the human IL20RA protein.

[0582] Preferably, all or part of the extracellular domain comprises the amino acid sequence shown in positions 30-250, 34-250, 30-247, or 34-247 of SEQ ID NO: 83; or, comprises an amino acid sequence with an identity of at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with ...10, 9, 8, 7, 6, 5, 4, 3, 2, or at least 1 amino acid with the amino acid sequence shown in positions 30-250, 34-250, 30-247, or 34-247 of SEQ ID NO: 83; or, comprises an amino acid sequence with an identity of at least 10, 9, 8, 7, 6, 5, 4, 3, 2, or at least 1 amino acid with the amino acid sequence shown in positions 30-250, 34-250, 30-247, or 34-247 of SEQ ID NO: 83; or, comprises an amino acid sequence with an identity of at least 10, 9, 8, 7, NO:83 indicates an amino acid sequence including substitutions, deletions, and / or insertions of one or more amino acid residues, as shown in positions 30-250, 34-250, 30-247, or 34-247.

[0583] Preferably, the non-human animal is a rodent, preferably a rat or mouse.

[0584] Preferably, the modified IL20RA protein contains a cytoplasmic domain that is substantially the same as that of the non-human animal endogenous IL20RA protein.

[0585] Preferably, the amino acid sequence of the cytoplasmic domain comprises the amino acid sequence shown in positions 275-546 of SEQ ID NO: 82; or, comprises an amino acid sequence with at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in positions 275-546 of SEQ ID NO: 82; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 275-546 of SEQ ID NO: 82 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with positions 275-546 of SEQ ID NO: 82, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0586] Preferably, the modified IL20RA protein contains a transmembrane domain that is substantially the same as that of the non-human endogenous IL20RA protein.

[0587] Preferably, the amino acid sequence of the transmembrane domain comprises the amino acid sequence shown in positions 254-274 of SEQ ID NO: 82; or, comprises an amino acid sequence with an identity of at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown in positions 254-274 of SEQ ID NO: 82; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 254-274 of SEQ ID NO: 82 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with an identity of positions 254-274 of SEQ ID NO: 82, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0588] Preferably, the modified IL20RA protein has an amino acid sequence comprising the amino acid sequence shown in SEQ ID NO: 91; or, having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in SEQ ID NO: 91; or, differing from the amino acid sequence shown in SEQ ID NO: 91 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, having an amino acid sequence similar to that shown in SEQ ID NO: 91, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0589] Preferably, the nucleotide sequence encoding the human IL20RA segment includes a portion of exon 2, all of exon 3, all of exon 4, all of exon 5, and a portion of exon 6 of the human IL20RA gene.

[0590] Preferably, the nucleotide sequence encoding the human IL20RA region comprises the nucleotide sequence shown in positions 137004735-137017092 of NCBI Gene Accession Number NC_000006.12; or, it comprises a nucleotide sequence with an identity of at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99. 8% or at least 99% of the nucleotide sequence; or, a nucleotide sequence that differs from the nucleotide sequence shown in positions 137004735-137017092 of NCBI Gene Accession Number NC_000006.12 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2 or no more than 1 nucleotide; or, a nucleotide sequence that includes substitutions, deletions and / or insertions of one or more nucleotides as shown in the nucleotide sequence shown in positions 137004735-137017092 of NCBI Gene Accession Number NC_000006.12.

[0591] Preferably, the mRNA transcribed from the nucleotide sequence encoding the modified IL20RA protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 90; B) the nucleotide sequence having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 90; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 90 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 90, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0592] Preferably, the expression of the modified IL20RA protein is controlled by endogenous IL20RA regulatory elements (e.g., promoter, 5'UTR and / or 3'UTR).

[0593] Preferably, the non-human animal or non-human animal genome is homozygous or heterozygous for the genetically modified endogenous IL20RA gene.

[0594] Preferably, the genome of the non-human animal or the non-human animal contains an endogenous gene encoding a human or modified other gene. Preferably, the other gene is selected from at least one of IL22, IL22RA1, IL10RB, IL20RB, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3, and OX40.

[0595] In a thirty-ninth aspect, the present invention provides a nucleic acid comprising a nucleotide sequence encoding an IL20RA protein modified as described above. Preferably, the nucleic acid further comprises the nucleotide sequence shown in positions 137004735-137017092 of NCBI Gene Accession Number NC_000006.12. Preferably, the mRNA transcribed from the nucleic acid comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 90; B) the identity with the nucleotide sequence shown in SEQ ID NO: 90 is at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99%; C) the difference from the nucleotide sequence shown in SEQ ID NO: 90 is no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 90, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0596] In a fortieth aspect of the present invention, a polypeptide is provided, said polypeptide being encoded by the genetically modified endogenous IL20RA gene described above or the nucleic acid described in the thirty-ninth aspect.

[0597] In a forty-first aspect, the present invention provides a genetically modified non-human animal or a method for constructing the thereof, wherein the genome of the non-human animal comprises at least one chromosome, the chromosome comprising a nucleotide sequence encoding a human or chimeric IL20RA protein (e.g., a humanized IL20RA protein). In some embodiments, the nucleotide sequence encoding the human or chimeric IL20RA protein may be a genomic DNA sequence, a CDS sequence, or a cDNA sequence. In some embodiments, the nucleotide sequence encoding the human or chimeric IL20RA protein is operatively linked to an endogenous regulatory element (e.g., a promoter, a 5'UTR, and / or a 3'UTR) at an endogenous IL20RA locus on at least one chromosome. In some embodiments, the chimeric IL20RA protein comprises a human or chimeric extracellular domain, a non-human animal endogenous transmembrane domain, and a non-human animal endogenous cytoplasmic domain. In some embodiments, the chimeric IL20RA protein comprises an endogenous signal peptide, a human or chimeric extracellular domain, a non-human animal endogenous transmembrane domain, and a non-human animal endogenous cytoplasmic domain. In some embodiments, the amino acid sequence of the human or chimeric IL20RA protein comprises SEQ ID NO: 83; or comprises an amino acid sequence with at least 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 99.5% identity to SEQ ID NO: 83. In some embodiments, the amino acid sequence of the human or chimeric IL20RA protein comprises the amino acid shown at positions 34-250, 30-250, 30-247, or 34-247 of SEQ ID NO: 83; or comprises an amino acid sequence with at least 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 99.5% identity to the amino acid shown at positions 34-250, 30-250, 30-247, or 34-247 of SEQ ID NO: 83. In some embodiments, the amino acid sequence of the human or chimeric IL20RA protein comprises SEQ ID NO: 91; or comprises an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 99.5% identical to SEQ ID NO: 91. In some embodiments, the non-human animal is a mammal, such as a monkey or a rodent (e.g., a mouse or rat). In some embodiments, the non-human animal is a mouse. In some embodiments, the endogenous IL20RA protein in the non-human animal is not expressed or is expressed at a reduced level compared to IL20RA in wild-type animals. In some embodiments, one or more cells of the non-human animal express the human or chimeric IL20RA protein.

[0598] In a forty-second aspect, the present invention provides a genetically modified non-human animal or a method thereof for construction, wherein in some embodiments, the genome of the non-human animal includes a nucleotide sequence at the endogenous IL20RA locus that replaces a nucleotide sequence encoding a region of human or chimeric IL20RA with a nucleotide sequence encoding a region of human or chimeric IL20RA. In some embodiments, the nucleotide sequence encoding the region of human or chimeric IL20RA is operatively linked to an endogenous regulatory element (e.g., an endogenous promoter, 5'UTR, and / or 3'UTR) at the endogenous IL20RA locus of the non-human animal. In some embodiments, one or more cells of the non-human animal express human or chimeric IL20RA protein (e.g., humanized IL20RA protein). In some embodiments, the endogenous IL20RA protein of the non-human animal is not expressed or is expressed at a reduced level compared to IL20RA in wild-type animals. In some embodiments, the replaced region encoding the region of endogenous IL20RA is all or part of the extracellular domain of the endogenous IL20RA. In some embodiments, one or more cells of the non-human animal express a chimeric IL20RA protein (e.g., a humanized IL20RA protein), the chimeric IL20RA comprising an extracellular domain, a transmembrane domain, and a cytoplasmic domain. In some embodiments, the extracellular domain is identical to or has at least 50%, 60%, 70%, 80%, 90%, 95%, or 99.9% identity with the extracellular domain of the human IL20RA protein. In some embodiments, the transmembrane domain and the cytoplasmic domain are identical to or have at least 50%, 60%, 70%, 80%, 90%, 95%, or 99.9% identity with the transmembrane domain and the cytoplasmic domain of the non-human animal endogenous IL20RA protein. In some embodiments, the extracellular domain is identical to the extracellular domain of the human IL20RA protein for at least 10, 20, 50, 100, 150, 200, 214, 217, 218, 220, or 221 consecutive amino acids. In some embodiments, the transmembrane domain and cytoplasmic domain are identical to the transmembrane domain and cytoplasmic domain of non-human endogenous IL20RA for at least 10, 20, 50, 100, 150, 200, 250, 290, or 293 consecutive amino acids (e.g., positions 254-546 of SEQ ID NO: 82). In some embodiments, the nucleotide sequence encoding the human or chimeric IL20RA region comprises all or part of exon 2 of the human IL20RA gene, all of exons 3 to 5, and part of exon 6. Preferably, the nucleotide sequence encoding the human or chimeric IL20RA region comprises at least 50-100, 100-1000, 1000-5000, or 10000-15000 bp of nucleotides from all or part of exon 2, all of exons 3 to 5, and part of exon 6 of the human IL20RA gene.In some embodiments, the nucleotide sequence encoding the human or chimeric IL20RA region comprises portions of exons 2 through 6 of the human IL20RA gene. In some embodiments, the nucleotide sequence encoding the corresponding region of human IL20RA is identical or identical to the nucleotide sequence shown at positions 137004735-137017092 of NCBI Gene Accession Number NC_000006.12 by at least 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 99.9%. In some embodiments, the nucleotide sequence of the replaced endogenous IL20RA region comprises portions of exons 2 through 6 of the non-human animal IL20RA gene. In some embodiments, the modified IL20RA gene in the animal genome is homozygous or heterozygous for the endogenously replaced locus.

[0599] In a forty-third aspect, the present invention provides a non-human animal or a method for constructing the thereof, wherein in some embodiments, the non-human animal comprises at least one cell encoding a nucleotide sequence of a human or chimeric IL20RA protein, wherein the chimeric IL20RA protein comprises a sequence of at least 50, 100, 200, 214, 217, 218, 220, 221, 300, 400, 500, 550, or 553 consecutive amino acids identical to the sequence of a human IL20RA protein. In some embodiments, the chimeric IL20RA protein comprises an amino acid sequence of at least 10, 20, 50, 100, 150, 200, 214, 217, 218, 220, or 221 consecutive amino acids identical to the sequence of an extracellular domain of human IL20RA. In some embodiments, the chimeric IL20RA protein comprises an amino acid sequence identical to at least 10, 20, 50, 100, 150, 200, 250, 290, or 293 consecutive amino acids of the transmembrane and cytoplasmic domains of nonhuman endogenous IL20RA. In some embodiments, the human or chimeric IL20RA protein comprises an amino acid sequence that is identical or has at least 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 99.9% identity with the amino acids shown at positions 34-250, 30-250, 30-247, or 34-247 of SEQ ID NO: 83. In some embodiments, the nucleotide sequence encoding the human or chimeric IL20RA protein is operatively linked to a regulatory element of nonhuman endogenous IL20RA (e.g., an endogenous promoter, 5'UTR, and / or 3'UTR). In some embodiments, the nucleotide sequence encoding the human or chimeric IL20RA protein may be integrated into the non-human animal endogenous IL20RA gene locus. In some embodiments, the human or chimeric IL20RA protein has at least one mouse IL20RA activity and / or human IL20RA activity.

[0600] In a forty-fourth aspect, the present invention provides a genetically modified non-human animal or a method thereof, wherein in some embodiments, in at least one cell of the non-human animal, at the non-human animal endogenous IL20RA gene locus, the nucleotide sequence encoding a region of non-human endogenous IL20RA is replaced by a nucleotide sequence encoding a region of human or chimeric IL20RA (or a nucleotide sequence encoding a human or chimeric IL20RA protein). In some embodiments, the endogenous IL20RA protein of the non-human animal is not expressed or is expressed at a reduced level compared to IL20RA in wild-type animals. Preferably, the relevant limitations on the nucleotide sequence encoding the region of human or chimeric IL20RA are the same as those described above. The relevant limitations on the amino acid sequence of the human or chimeric IL20RA protein are the same as those described above. In some embodiments, the nucleotide sequence encoding the region of non-human endogenous IL20RA comprises portions of exons 2 to 6 of the non-human animal IL20RA gene. In some embodiments, the nucleotide sequence encoding the region of human or chimeric IL20RA is operatively linked to a regulatory element of the non-human endogenous IL20RA (e.g., a promoter, 5'UTR, and / or 3'UTR). In some embodiments, the non-human animal is a mammal, such as a monkey or a rodent (e.g., a mouse or a rat). In some embodiments, the non-human animal is a mouse.

[0601] In a forty-fifth aspect, the present invention provides a method for constructing a gene-modified non-human animal cell expressing human or chimeric IL20RA, the method comprising replacing, at the non-human animal endogenous IL20RA gene locus, a nucleotide sequence encoding an endogenous IL20RA region with a nucleotide sequence encoding a human or chimeric IL20RA protein (or a nucleotide sequence encoding a human or chimeric IL20RA protein), thereby generating a gene-modified non-human animal cell expressing the human or chimeric IL20RA protein. Preferably, the relevant limitations on the nucleotide sequence encoding the human or chimeric IL20RA region are the same as those described above. The relevant limitations on the amino acid sequence of the human or chimeric IL20RA protein are the same as those described above. In some embodiments, the replaced endogenous IL20RA region is all or part of the extracellular domain of the non-human animal endogenous IL20RA. In some embodiments, the replaced nucleotide sequence encoding the endogenous IL20RA region comprises portions of exons 2 to 6 of the non-human animal IL20RA gene. In some embodiments, the nucleotide sequence encoding the human or chimeric IL20RA protein is operatively linked to a regulatory element of endogenous IL20RA (e.g., a promoter, 5'UTR, and / or 3'UTR). In some embodiments, the non-human animal is a mouse. In some embodiments, the non-human animal further includes nucleotide sequences of other genes encoding human or chimeric proteins selected from at least one of IL22, IL22RA1, IL10RB, IL20RB, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3, and OX40. Preferably, the IL22 is a chimeric IL22 protein; more preferably, the chimeric IL22 protein comprises any of the humanized IL22 proteins described in this invention. Preferably, the IL22RA1 is a chimeric IL22RA1 protein; more preferably, the chimeric IL22RA1 protein comprises any of the humanized IL22RA1 proteins described in this invention. Preferably, the IL10RB is a chimeric IL10RB protein; more preferably, the chimeric IL10RB protein comprises any of the humanized IL10RB proteins described in this invention. Preferably, the IL20RB is a chimeric IL20RB protein; more preferably, the chimeric IL20RB protein comprises any of the humanized IL20RB proteins described in this invention.

[0602] In a forty-sixth aspect, the present invention provides a humanized IL20RA protein, wherein in some embodiments, the humanized IL20RA protein comprises all or part of the human IL20RA protein. In some embodiments, the humanized IL20RA protein comprises an amino acid sequence that is identical or identical in at least 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 99.9% to the amino acid sequence shown in SEQ ID NO: 83, SEQ ID NO: 83 positions 34-250, SEQ ID NO: 83 positions 30-250, SEQ ID NO: 83 positions 30-247, and SEQ ID NO: 83 positions 34-247. In some embodiments, the humanized IL20RA protein comprises an amino acid sequence that is identical or identical in at least 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 99.9% to the amino acid sequence shown in SEQ ID NO: 91.

[0603] In a forty-seventh aspect, the present invention provides a humanized IL20RA gene, wherein in some embodiments, the humanized IL20RA gene encodes the humanized IL20RA protein. In some embodiments, the humanized IL20RA gene comprises all or a portion of exon 2 of the human IL20RA gene, all of exons 3 to 5, and a portion of exon 6. In some embodiments, the humanized IL20RA gene comprises portions of exons 2 to 6 of the human IL20RA gene. The portion of exon 2 comprises at least a 20 bp continuous nucleotide sequence, for example, at least 20, 30, 40, 50, 60, 70, 80, 90, 100, 110, 120, 125, 130, or 136 bp continuous nucleotide sequences, preferably comprising all or a portion of the nucleotide sequence encoding the extracellular domain in exon 2. The exon 6 portion comprises at least a 5 bp continuous nucleotide sequence, such as 5, 10, 15, 20, 21, 22, 23, 24, 25, 26, 30, 40, 50, 60, 70, 80, 90, 100, 110, 120, 130, or 140 bp continuous nucleotide sequences, preferably comprising all or part of the nucleotide sequence encoding the extracellular domain in exon 6. In some embodiments, the humanized IL20RA gene comprises the entire exon 1, a portion of exon 2, a portion of exon 6, and the entire exon 7 of the non-human IL20RA gene. In some embodiments, the humanized IL20RA gene comprises the entire exon 1, a portion of exon 6, and the entire exon 7 of the non-human IL20RA gene. In some embodiments, the nucleotide sequence comprising the humanized IL20RA gene is identical or identical to the nucleotide sequence shown in SEQ ID NO: 84, 85, 86, 87, 88, 89, 90, 92, 93 or NCBI Gene Accession Number NC_000006.12 at positions 137004735-137017092 by at least 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 99.9%.

[0604] In a forty-eighth aspect of the present invention, a targeting vector is provided, comprising: a) a 5' homologous DNA fragment (5' arm or 5' homologous arm) of the conversion region to be modified, selected from the genomic DNA of the IL20RA gene, having a length of 100 to 10,000 nucleotides; b) a donor region; and c) a 3' homologous DNA fragment (5' arm or 5' homologous arm) of the conversion region to be modified, selected from the genomic DNA of the IL20RA gene, having a length of 100 to 10,000 nucleotides.

[0605] In some embodiments, a) the DNA fragment homologous to the 5' end of the transition region to be modified is selected from a nucleotide sequence with at least 90% homology to the NCBI accession number NC_000076.7; a) the DNA fragment homologous to the 3' end of the transition region to be modified is selected from a nucleotide sequence with at least 90% homology to the NCBI accession number NC_000076.7.

[0606] In some embodiments, the length of the genomic nucleotide sequence selected for the targeting vector can exceed 0.8kb, 1kb, 2kb, 3kb, 4kb, 5kb, 6kb, 7kb, 8kb, 9kb, 10kb, 15kb, 16kb, 17kb, 18kb, 19kb, or 20kb.

[0607] In some embodiments, the conversion region to be altered is located on exons 1 to 7 of the non-human animal endogenous IL20RA gene, preferably on exons 2 to 6.

[0608] In some embodiments, the 5' homologous arm comprises SEQ ID NO: 84 and / or 92.

[0609] In some embodiments, the 3' homologous arm comprises SEQ ID NO: 85 and / or 93.

[0610] In some embodiments, the donor sequence comprises a human sequence, such as bits 137004735-137017092 of NCBI Gene Accession Number NC_000006.12.

[0611] In some embodiments, the targeting vector comprises one or more marker genes (or resistance genes), such as resistance genes for positive clone selection or genes encoding negative selection markers. In some embodiments, the resistance gene for positive clone selection is the neomycin phosphotransferase coding sequence Neo. Preferably, the targeting vector further comprises two co-directed Frt recombination sites flanking the marker gene. In some embodiments, the gene encoding the negative selection marker comprises the gene encoding the diphtheria toxin A subunit (DTA).

[0612] This invention also provides vectors for constructing humanized animal models or knockout models. In some embodiments, the vector contains an sgRNA sequence that targets the IL20RA gene. In some embodiments, the target site of the sgRNA in the conversion region to be altered is unique and satisfies the sequence arrangement rule of 5'-NNN(20)-NGG3' or 5'-CCN-N(20)-3'. In some embodiments, the target site of the sgRNA targeting the non-human IL20RA gene is located on exon 2 and / or exon 6 of the IL20RA gene.

[0613] Preferably, the target site sequence of the sgRNA targeting the IL20RA gene in non-human animals includes SEQ ID NO: 94 and / or 95.

[0614] Preferably, the sgRNA targeting the non-human animal IL20RA gene includes nucleotide sequences as shown in SEQ ID NO: 96 and 98, or nucleotide sequences as shown in SEQ ID NO: 97 and 99, or nucleotide sequences as shown in SEQ ID NO: 100 and 102, or nucleotide sequences as shown in SEQ ID NO: 101 and 103.

[0615] In some embodiments, the present invention relates to plasmid constructs containing sgRNA (sgRNA vectors, such as pT7-sgRNA vectors) and / or cells containing said constructs.

[0616] The present invention also relates to cells comprising the targeting vector or sgRNA vector as described above.

[0617] In some embodiments, the present invention also provides a non-human mammalian cell having any of the above-described targeting vectors and one or more in vitro transcripts from the plasmid construction system described in this application. In some embodiments, the non-human mammalian cell contains Cas9 mRNA or its in vitro transcript.

[0618] In some embodiments, the genes in the non-human mammalian cells are heterozygous. In some embodiments, the genes in the non-human mammalian cells are homozygous.

[0619] In some embodiments, the non-human mammalian cell is a mouse cell. In some embodiments, the non-human mammalian cell is a fertilized egg cell. In some embodiments, the non-human mammalian cell is an embryonic stem cell. In some embodiments, the non-human mammalian cell is any type of cell capable of expressing IL20RA.

[0620] In a forty-ninth aspect of the present invention, a non-human animal genome is provided, wherein the non-human animal or the non-human animal genome comprises a genetically modified endogenous IL20RB gene encoding a modified IL20RB protein, and the non-human animal expresses the modified IL20RB protein.

[0621] Preferably, the non-human animal or non-human animal genome contains a nucleotide sequence encoding the corresponding segment of the non-human animal endogenous IL20RB, which is replaced by a nucleotide sequence encoding the human IL20RB segment.

[0622] Preferably, the modified IL20RB protein comprises a cytoplasmic domain, a transmembrane domain, and an extracellular domain.

[0623] Preferably, the modified IL20RB protein contains an extracellular domain substantially the same as that of the human IL20RB protein.

[0624] Preferably, all or part of the extracellular domain comprises the amino acid sequence shown in positions 44-233, 30-233, or 35-233 of SEQ ID NO: 113; or, comprises an amino acid sequence with an identity of at least 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with ...10, 9, 8, 7, 6, 5, 4, 3, 2, or at least 1 amino acid with the amino acid sequence shown in positions 44-233, 30-233, or 35-233 of SEQ ID NO: 113; or, comprises an amino acid sequence with an identity of at least 10, 9, 8, 7, 6, 5, 4, 3, 2, or at least 1 amino acid with the amino acid sequence shown in positions 44-233, 30-233, or 35-233 of SEQ ID NO: 113; or, comprises an amino acid sequence with an identity of at least 10, 9, 8, 7, 6, 5, 4, 3, 2, or at least 1 amino acid with the amino acid sequence shown The amino acid sequence shown in NO:113 at positions 44-233, 30-233, or 35-233, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0625] Preferably, the extracellular domain comprises 0-15 amino acid residues (e.g., 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15) at the N-terminus and / or C-terminus of the extracellular domain of the non-human endogenous IL20RB protein, more preferably comprising the amino acid sequence shown at positions 27-38 of SEQ ID NO:112; or, comprising amino acid sequences with at least 77%, 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown at positions 27-38 of SEQ ID NO:112; or, comprising amino acid sequences with differences of no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid from ... The amino acid sequence shown at positions 27-38 of NO:112 includes substitutions, deletions, and / or insertions of one or more amino acid residues.

[0626] Preferably, the modified IL20RB protein contains a transmembrane domain that is substantially the same as that of the human IL20RB protein.

[0627] Preferably, all or part of the transmembrane domain comprises the amino acid sequence shown at positions 234-250, 234-244, or 234-254 of SEQ ID NO: 113; or, comprises an amino acid sequence with an identity of at least 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with ...10, 9, 8, 7, 6, 5, 4, 3, 2, or at least 1 amino acid with the amino acid sequence shown at positions 234-250, 234-244, or 234-254 of SEQ ID NO: 113; or, comprises an amino acid sequence with an identity of at least 10, 9, 8, 7, 6, 5, 4, 3, 2, or at least 1 amino acid with the amino acid sequence shown at positions 234-250, 234-244, or 234-254 of SEQ ID NO: 113; or, comprises an amino acid sequence with an identity of at least 10, 9, 8, 7, 6, 5, 4, NO: 113 indicates the amino acid sequence represented by positions 234-250, 234-244, or 234-254, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0628] Preferably, the transmembrane domain comprises 0-15 amino acid residues (e.g., 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15) at the N-terminus and / or C-terminus of the transmembrane domain of a non-human endogenous IL20RB protein, more preferably comprising the amino acid sequence shown at positions 240-249 of SEQ ID NO:112; or, comprising amino acid sequences with at least 77%, 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown at positions 240-249 of SEQ ID NO:112; or, comprising amino acid sequences with at least 99% identity with the amino acid sequence shown at positions 240-249 of SEQ ID NO:112; or, comprising amino acid sequences with at least 99% identity with the amino acid sequence shown at positions 240-249 of SEQ ID NO:112. The amino acid sequence shown at positions 240-249 of SEQ ID NO:112 differs by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2 or no more than 1 amino acid; or, it contains an amino acid sequence that is identical to the amino acid sequence shown at positions 240-249 of SEQ ID NO:112, including substitutions, deletions and / or insertions of one or more amino acid residues.

[0629] Preferably, the non-human animal is a rodent, preferably a rat or mouse.

[0630] Preferably, the modified IL20RB protein contains a cytoplasmic domain that is substantially the same as that of the non-human endogenous IL20RB protein.

[0631] Preferably, the amino acid sequence of the cytoplasmic domain comprises the amino acid sequence shown in positions 250-308 of SEQ ID NO: 112; or, comprises an amino acid sequence with at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in positions 250-308 of SEQ ID NO: 112; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 250-308 of SEQ ID NO: 112 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with positions 250-308 of SEQ ID NO: 112, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0632] Preferably, the modified IL20RB protein has an amino acid sequence comprising the amino acid sequence shown in SEQ ID NO: 121; or, having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in SEQ ID NO: 121; or, differing from the amino acid sequence shown in SEQ ID NO: 121 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, having an amino acid sequence similar to that shown in SEQ ID NO: 121, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0633] Preferably, the nucleotide sequence encoding the human IL20RB region comprises portions of exons 2 to 6 of the human IL20RB gene, wherein the portion of exon 2 comprises at least 20 bp of continuous nucleotide sequence, such as at least 20, 30, 40, 50, 60, 70, 80, 85, 86, 90, 100, 110, 120, or 127 bp of continuous nucleotide sequence. The portion of exon 6 comprises at least 20 bp of continuous nucleotide sequence, such as at least 20, 30, 40, 50, 60, 70, 80, 85, 86, 90, 100, 110, 120, 130, 140, or 143 bp of continuous nucleotide sequence.

[0634] Preferably, the nucleotide sequence encoding the human IL20RB region comprises the nucleotide sequence shown in positions 136980507-136995463 of NCBI Gene Accession Number NC_000003.12; or, it comprises a nucleotide sequence with an identity of at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 9 ... 8% or at least 99% of the nucleotide sequence; or, a nucleotide sequence that differs from the nucleotide sequence shown in positions 136980507-136995463 of NCBI Gene Accession Number NC_000003.12 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2 or no more than 1 nucleotide; or, a nucleotide sequence that includes substitutions, deletions and / or insertions of one or more nucleotides as shown in the nucleotide sequence shown in positions 136980507-136995463 of NCBI Gene Accession Number NC_000003.12.

[0635] Preferably, the mRNA transcribed from the nucleotide sequence encoding the modified IL20RB protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 120; B) the nucleotide sequence having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 120; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 120 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 120, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0636] Preferably, the nucleotide sequence of the replaced segment encoding the corresponding region of non-human endogenous IL20RB comprises all or part of the nucleotide sequence encoding the extracellular domain of the non-human endogenous IL20RB protein, more preferably comprising the nucleotide sequence encoding positions 27-225 or 39-225 of SEQ ID NO: 112, or comprising a nucleotide sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence encoding positions 27-225 or 39-225 of SEQ ID NO: 112; or comprising a nucleotide sequence with a difference of no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide from ... of NO: 112 The nucleotide sequence shown at positions 27-225 or 39-225 includes a nucleotide sequence with substitutions, deletions and / or insertions of one or more nucleotides, more preferably a partial nucleotide sequence containing exons 2 to 6 of a non-human animal endogenous IL20RB sequence.

[0637] Preferably, the nucleotide sequence of the replaced segment encoding the corresponding region of non-human endogenous IL20RB comprises all or part of the nucleotide sequence encoding the transmembrane domain of the non-human endogenous IL20RB protein, more preferably comprising the nucleotide sequence encoding positions 226-239 or 226-249 of SEQ ID NO: 112, or comprising a nucleotide sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity to the nucleotide sequence encoding positions 226-239 or 226-249 of SEQ ID NO: 112; or comprising a nucleotide sequence encoding SEQ ID NO: 112 with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity to the nucleotide sequence encoding SEQ ID NO: 112; or comprising a nucleotide sequence encoding SEQ ID NO: 112 with at least 99% identity to the nucleotide sequence encoding SEQ ID NO: 112. The nucleotide sequence difference between positions 226-239 or 226-249 of SEQ ID NO: 112 is no more than 10, 9, 8, 7, 6, 5, 4, 3, 2 or no more than 1 nucleotide; or, it contains a nucleotide sequence that is identical to the nucleotide sequence encoding positions 226-239 or 226-249 of SEQ ID NO: 112, including substitutions, deletions and / or insertions of one or more nucleotides, more preferably a partial nucleotide sequence containing exon 6 of a non-human animal endogenous IL20RB sequence.

[0638] Preferably, the non-human animal genome includes replacing all or part of the nucleotide sequence encoding the extracellular domain of the endogenous IL20RB protein in the non-human animal genome with a nucleotide sequence encoding an extracellular domain substantially identical to that of the human IL20RB protein.

[0639] Preferably, the non-human animal genome includes replacing all or part of the nucleotide sequence encoding the transmembrane domain of the endogenous IL20RB protein in the non-human animal genome with a nucleotide sequence encoding a transmembrane domain substantially identical to that of the human IL20RB protein.

[0640] Preferably, the non-human animal genome includes replacing all or part of the nucleotide sequences encoding the extracellular domain and transmembrane domain of the endogenous IL20RB protein in the non-human animal genome with nucleotide sequences encoding the extracellular domain and transmembrane domain that are substantially identical to those of the human IL20RB protein.

[0641] Preferably, the non-human animal genome includes replacing all or part of the nucleotide sequence encoding the extracellular domain of the endogenous IL20RB protein in the non-human animal genome with a nucleotide sequence encoding the human IL20RB region.

[0642] Preferably, the non-human animal genome includes replacing all or part of the nucleotide sequence encoding the transmembrane domain of the endogenous IL20RB protein in the non-human animal genome with the nucleotide sequence encoding the human IL20RB region.

[0643] Preferably, the non-human animal genome includes replacing all or part of the extracellular domain and transmembrane domain of the non-human animal genome encoding the endogenous IL20RB protein with the nucleotide sequence encoding the human IL20RB region.

[0644] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding positions 39-225 or 27-225 of SEQ ID NO: 112 in the non-human animal genome with a nucleotide sequence encoding the human IL20RB region (e.g., genomic DNA sequence, cDNA sequence or CDS sequence).

[0645] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding positions 226-249 or 226-239 of SEQ ID NO: 112 in the non-human animal genome with a nucleotide sequence encoding the human IL20RB region (e.g., genomic DNA sequence, cDNA sequence or CDS sequence).

[0646] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding positions 39-239 or 27-249 of SEQ ID NO: 112 in the non-human animal genome with a nucleotide sequence encoding the human IL20RB region (e.g., genomic DNA sequence, cDNA sequence, or CDS sequence).

[0647] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding positions 39-225 or 27-225 of SEQ ID NO: 112 in the non-human animal genome with a portion of exon 2 to exon 6 of the human IL20RB gene.

[0648] Preferably, the non-human animal genome includes replacing parts of exons 2 to 6 of the non-human animal endogenous IL20RB gene in the non-human animal genome with parts of exons 2 to 6 of the human IL20RB gene.

[0649] Preferably, the non-human animal genome includes replacing part of exon 6 of the human IL20RB gene with the nucleotide sequence encoding positions 226-239 or 226-249 of SEQ ID NO: 112 in the non-human animal genome.

[0650] Preferably, the non-human animal genome includes replacing part of exon 6 of the non-human animal endogenous IL20RB gene in the non-human animal genome with part of exon 6 of the human IL20RB gene.

[0651] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding positions 39-239 or 27-225 of SEQ ID NO: 112 with portions of exons 2 to 6 of the human IL20RB gene.

[0652] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding positions 39-239 of SEQ ID NO: 112 in the non-human animal genome with the nucleotide sequence encoding positions 44-244 of SEQ ID NO: 113.

[0653] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding positions 27-249 of SEQ ID NO: 112 in the non-human animal genome with the nucleotide sequence encoding positions 30-254 of SEQ ID NO: 113.

[0654] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding SEQ ID NO: 112 in the non-human animal genome with the nucleotide sequence encoding SEQ ID NO: 121.

[0655] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding positions 39-239 of SEQ ID NO: 112 in the non-human animal genome with the nucleotide sequence shown in positions 136980507-136995463 of NCBI accession number NC_000003.12.

[0656] Preferably, the non-human animal genome includes replacing the nucleotide sequence encoding SEQ ID NO: 112 in the non-human animal genome with the nucleotide sequence shown in SEQ ID NO: 120.

[0657] Preferably, the expression of the modified IL20RB protein is controlled by endogenous IL20RB regulatory elements (e.g., promoter, 5'UTR and / or 3'UTR).

[0658] Preferably, the non-human animal genome is homozygous or heterozygous for the genetically modified endogenous IL20RB gene.

[0659] Preferably, the non-human animal genome contains endogenous genes encoding human or modified or wild-type other genes, and preferably, the other genes are selected from at least one of IL22, IL22RA1, IL10RB, IL20RA, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3 and OX40.

[0660] Preferably, the non-human animal genome contains an endogenous IL10RB locus encoding a human, modified, or wild-type IL10RB protein.

[0661] In one specific embodiment of the present invention, the non-human animal genome contains a genetically modified endogenous IL10RB locus encoding a modified IL10RB protein, and the non-human animal expresses the modified IL10RB protein.

[0662] Preferably, in the genetically modified endogenous IL10RB locus, the nucleotide sequence encoding the corresponding segment of non-human endogenous IL10RB is replaced with the nucleotide sequence encoding the human IL10RB segment.

[0663] Preferably, the modified IL10RB protein comprises the amino acid sequence shown in SEQ ID NO: 62; or, comprises an amino acid sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in SEQ ID NO: 62; or, comprises an amino acid sequence differing from the amino acid sequence shown in SEQ ID NO: 62 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence similar to that shown in SEQ ID NO: 62, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0664] Preferably, the nucleotide sequence encoding the human IL10RB region includes a portion of the human IL10RB gene; more preferably, it includes a portion of exon 2, all of exon 3, all of exon 4, all of exon 5, and a portion of exon 6 of the human IL10RB gene; even more preferably, it includes portions of exons 2 to 6.

[0665] Preferably, the nucleotide sequence encoding the human IL10RB region comprises the nucleotide sequence shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or a variant thereof; or, comprises a nucleotide sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity to the nucleotide sequence shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or a variant thereof; or, comprises a nucleotide sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity to the nucleotide sequence shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or a variant thereof. The nucleotide sequence shown at positions 33268405-33288111 or its variants differ by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2 or no more than 1 nucleotide; or, a nucleotide sequence containing the nucleotide sequence shown at positions 33268405-33288111 or its variants with NCBI gene accession number NC_000021.9, including substitutions, deletions and / or insertions of one or more nucleotides, wherein the variants include one or more of the following: C>T mutation at position 33276503, G>A mutation at position 33276818, or TTTTGGG>TTG mutation at positions 33277048-33277053;

[0666] Preferably, the mRNA transcribed from the nucleotide sequence encoding the modified IL10RB protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 61; B) the nucleotide sequence having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 61; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 61 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 61, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0667] Preferably, the non-human animal genome contains an endogenous IL10 locus encoding a human, modified, or wild-type IL10 protein.

[0668] Preferably, the non-human animal or non-human animal genome contains a genetically modified endogenous IL10 locus encoding a human or modified IL10 protein, and the non-human animal expresses the human or modified IL10 protein.

[0669] Preferably, in the genetically modified endogenous IL10 locus, the nucleotide sequence of the corresponding segment of the non-human animal endogenous IL10 is replaced with the nucleotide sequence encoding the human IL10 segment.

[0670] Preferably, the nucleotide sequence encoding the human IL10 region comprises a portion of exon 1, all of exon 2, all of exon 3, all of exon 4, and a portion of exon 5 of the human IL10 gene; more preferably, it comprises portions of exons 1 to 5. The portion of exon 1 comprises at least 50 bp, for example, at least 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, 160, 165, 170, 180, 190, 200, 220, or 224 bp of continuous nucleotide sequence. The portion of exon 5 contains at least 50 bp, such as at least 50, 60, 70, 80, 90, 93, 100, 200, 300, 400, 500, 600, 700, 800, 900, 1000, 1100, 1120, or 1127 bp of continuous nucleotide sequence.

[0671] Preferably, the non-human animal genome contains an endogenous IL10RA locus encoding a human, modified, or wild-type IL10RA protein.

[0672] Preferably, the non-human animal or the non-human animal genome contains a genetically modified endogenous IL10RA locus encoding a modified IL10RA protein, and the non-human animal expresses the modified IL10RA protein.

[0673] Preferably, in the genetically modified endogenous IL10RA locus, the nucleotide sequence encoding the corresponding segment of non-human animal endogenous IL10RA is replaced with the nucleotide sequence encoding the human IL10RA segment.

[0674] Preferably, the modified IL10RA protein comprises a cytoplasmic domain, a transmembrane domain, and an extracellular domain.

[0675] Preferably, the modified IL10RA protein further comprises a signal peptide.

[0676] Preferably, the IL10RA protein contains a signal peptide that is substantially the same as that of the human IL10RA protein.

[0677] Preferably, the IL10RA protein contains an extracellular domain that is substantially the same as that of the human IL10RA protein.

[0678] Preferably, the IL10RA protein contains a transmembrane domain that is substantially the same as that of the human IL10RA protein.

[0679] Preferably, the IL10RA protein contains a cytoplasmic domain that is substantially the same as that of the human IL10RA protein.

[0680] Preferably, the cytoplasmic domain also includes all or part of the cytoplasmic domain of the non-human animal endogenous IL10RA protein.

[0681] Preferably, the nucleotide sequence encoding the human IL10RA segment includes a portion of exon 1, all of exon 2, all of exon 3, all of exon 4, all of exon 5, and a portion of exon 6 of the human IL10RA gene; more preferably, it includes portions of exons 1 to 6.

[0682] Preferably, the non-human animal genome comprises any one of the following groups:

[0683] 1) Genetically modified endogenous IL20RB gene and wild-type endogenous IL20RA gene encoding modified IL20RB protein;

[0684] 2) Genetically modified endogenous IL20RB gene, wild-type endogenous IL20RA gene, and wild-type endogenous IL19 gene encoding modified IL20RB protein;

[0685] 3) Genetically modified endogenous IL20RB gene encoding modified IL20RB protein, genetically modified endogenous IL20RA gene encoding modified IL20RA protein, and wild-type endogenous IL19 gene;

[0686] 4) Genetically modified endogenous IL20RB gene, wild-type endogenous IL20RA gene, and wild-type endogenous IL20 gene encoding modified IL20RB protein;

[0687] 5) Genetically modified endogenous IL20RB gene encoding modified IL20RB protein, genetically modified endogenous IL20RA gene encoding modified IL20RA protein, and wild-type endogenous IL20 gene;

[0688] 6) Genetically modified endogenous IL20RB gene, wild-type endogenous IL20RA gene, and wild-type endogenous IL24 gene encoding modified IL20RB protein;

[0689] 7) Genetically modified endogenous IL20RB gene encoding modified IL20RB protein, genetically modified endogenous IL20RA gene encoding modified IL20RA protein, and wild-type endogenous IL24 gene;

[0690] 8) Genetically modified endogenous IL20RB gene and wild-type endogenous IL22RA1 gene encoding modified IL20RB protein;

[0691] 9) Genetically modified endogenous IL20RB gene, wild-type endogenous IL22RA1 gene and wild-type endogenous IL20 gene encoding modified IL20RB protein;

[0692] 10) Genetically modified endogenous IL20RB gene encoding modified IL20RB protein, genetically modified endogenous IL22RA1 gene encoding modified IL22RA1 protein, and wild-type endogenous IL20 gene;

[0693] 11) Genetically modified endogenous IL20RB gene, wild-type endogenous IL22RA1 gene and wild-type endogenous IL24 gene encoding modified IL20RB protein;

[0694] 12) Genetically modified endogenous IL20RB gene encoding modified IL20RB protein, genetically modified endogenous IL22RA1 gene encoding modified IL22RA1 protein, and wild-type endogenous IL24 gene.

[0695] According to a fiftieth aspect of the present invention, a non-human animal is provided, said non-human animal comprising a genetically modified endogenous IL20RB gene encoding a modified IL20RB protein, said non-human animal expressing said modified IL20RB protein.

[0696] The forty-ninth aspect of the present invention relates to limitations on modified IL20RB proteins and genetically modified endogenous IL20RB genes.

[0697] Preferably, the non-human animal comprises the non-human animal genome described in aspect forty-nine.

[0698] In a fifty-first aspect, the present invention provides a method for preparing a non-human animal, wherein the non-human animal or the genome of the non-human animal comprises a genetically modified endogenous IL20RB gene encoding a modified IL20RB protein, and the non-human animal expresses the modified IL20RB protein.

[0699] Preferably, the genome of the non-human animal or non-human animal contains a nucleotide sequence that replaces the nucleotide sequence encoding the corresponding segment of the non-human animal endogenous IL20RB with a nucleotide sequence encoding the human IL20RB segment.

[0700] Preferably, the modified IL20RB protein comprises a cytoplasmic domain, a transmembrane domain, and an extracellular domain.

[0701] Preferably, the modified IL20RB protein contains an extracellular domain substantially the same as that of the human IL20RB protein.

[0702] Preferably, all or part of the extracellular domain comprises the amino acid sequence shown in positions 44-233, 30-233, or 35-233 of SEQ ID NO: 113; or, comprises an amino acid sequence with an identity of at least 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with ...10, 9, 8, 7, 6, 5, 4, 3, 2, or at least 1 amino acid with the amino acid sequence shown in positions 44-233, 30-233, or 35-233 of SEQ ID NO: 113; or, comprises an amino acid sequence with an identity of at least 10, 9, 8, 7, 6, 5, 4, 3, 2, or at least 1 amino acid with the amino acid sequence shown in positions 44-233, 30-233, or 35-233 of SEQ ID NO: 113; or, comprises an amino acid sequence with an identity of at least 10, 9, 8, 7, 6, 5, 4, 3, 2, or at least 1 amino acid with the amino acid sequence shown The amino acid sequence shown in NO:113 at positions 44-233, 30-233, or 35-233, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0703] Preferably, the modified IL20RB protein contains a transmembrane domain that is substantially the same as that of the human IL20RB protein.

[0704] Preferably, all or part of the transmembrane domain comprises the amino acid sequence shown at positions 234-250, 234-244, or 234-254 of SEQ ID NO: 113; or, comprises an amino acid sequence with an identity of at least 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with ...10, 9, 8, 7, 6, 5, 4, 3, 2, or at least 1 amino acid with the amino acid sequence shown at positions 234-250, 234-244, or 234-254 of SEQ ID NO: 113; or, comprises an amino acid sequence with an identity of at least 10, 9, 8, 7, 6, 5, 4, 3, 2, or at least 1 amino acid with the amino acid sequence shown at positions 234-250, 234-244, or 234-254 of SEQ ID NO: 113; or, comprises an amino acid sequence with an identity of at least 10, 9, 8, 7, 6, 5, 4, NO: 113 indicates the amino acid sequence represented by positions 234-250, 234-244, or 234-254, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0705] Preferably, the nucleotide sequence encoding the human IL20RB region comprises the amino acid sequence shown in positions 44-244 of SEQ ID NO: 113; or, comprises an amino acid sequence with an identity of at least 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown in positions 44-244 of SEQ ID NO: 113; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 44-244 of SEQ ID NO: 113 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with an identity of positions 44-244 of SEQ ID NO: 113, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0706] Preferably, the non-human animal is a rodent, preferably a rat or mouse;

[0707] Preferably, the modified IL20RB protein contains a cytoplasmic domain that is substantially the same as that of the non-human endogenous IL20RB protein.

[0708] Preferably, the amino acid sequence of the cytoplasmic domain comprises the amino acid sequence shown in positions 250-308 of SEQ ID NO: 112; or, comprises an amino acid sequence with at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in positions 250-308 of SEQ ID NO: 112; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 250-308 of SEQ ID NO: 112 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with positions 250-308 of SEQ ID NO: 112, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0709] Preferably, the modified IL20RB protein has an amino acid sequence comprising the amino acid sequence shown in SEQ ID NO: 121; or, having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in SEQ ID NO: 121; or, differing from the amino acid sequence shown in SEQ ID NO: 121 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, having an amino acid sequence similar to that shown in SEQ ID NO: 121, including substitutions, deletions, and / or insertions of one or more amino acid residues.

[0710] Preferably, the nucleotide sequence encoding the human IL20RB region includes a portion of exon 2, all of exon 3, all of exon 4, all of exon 5, and a portion of exon 6 of the human IL20RB gene.

[0711] Preferably, the nucleotide sequence encoding the human IL20RB region comprises the nucleotide sequence shown in positions 136980507-136995463 of NCBI Gene Accession Number NC_000003.12; or, it comprises a nucleotide sequence with an identity of at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 9 ... 8% or at least 99% of the nucleotide sequence; or, a nucleotide sequence that differs from the nucleotide sequence shown in positions 136980507-136995463 of NCBI Gene Accession Number NC_000003.12 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2 or no more than 1 nucleotide; or, a nucleotide sequence that includes substitutions, deletions and / or insertions of one or more nucleotides as shown in the nucleotide sequence shown in positions 136980507-136995463 of NCBI Gene Accession Number NC_000003.12.

[0712] Preferably, the mRNA transcribed from the nucleotide sequence encoding the modified IL20RB protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 120; B) the nucleotide sequence having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 120; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 120 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 120, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0713] Preferably, the expression of the modified IL20RB protein is controlled by endogenous IL20RB regulatory elements (e.g., promoter, 5'UTR and / or 3'UTR).

[0714] Preferably, the non-human animal or non-human animal genome is homozygous or heterozygous for the genetically modified endogenous IL20RB gene.

[0715] Preferably, the genome of the non-human animal or the non-human animal contains an endogenous gene encoding a human or modified other gene. Preferably, the other gene is selected from at least one of IL22, IL22RA1, IL10RB, IL20RA, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3, and OX40.

[0716] In a fifty-second aspect, the present invention provides a nucleic acid comprising a nucleotide sequence encoding an IL20RB protein modified as described above. Preferably, the nucleic acid further comprises the nucleotide sequence shown in positions 136980507-136995463 of NCBI accession number NC_000003.12. Preferably, the mRNA transcribed from the nucleic acid comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 120; B) the identity with the nucleotide sequence shown in SEQ ID NO: 120 is at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99%; C) the difference from the nucleotide sequence shown in SEQ ID NO: 120 is no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 120, including substitutions, deletions, and / or insertions of one or more nucleotides.

[0717] In a fifty-third aspect, the present invention provides a polypeptide encoded by the genetically modified endogenous IL20RB gene described above or the nucleic acid described in aspect fifty-two.

[0718] In a fifty-fourth aspect, the present invention provides a genetically modified non-human animal or a method for constructing the same, wherein the genome of the non-human animal comprises at least one chromosome, the chromosome comprising a nucleotide sequence encoding a human or chimeric IL20RB protein. In some embodiments, the nucleotide sequence encoding the human or chimeric IL20RB protein is operatively linked to an endogenous regulatory element (e.g., a promoter, 5'UTR, and / or 3'UTR) at an endogenous IL20RB locus on at least one chromosome. In some embodiments, the chimeric IL20RB protein comprises a chimeric extracellular domain, a chimeric transmembrane domain, and a non-human animal endogenous cytoplasmic domain. In some embodiments, the chimeric IL20RB protein comprises an endogenous signal peptide, a chimeric extracellular domain, a chimeric transmembrane domain, and a non-human animal endogenous cytoplasmic domain. In some embodiments, the amino acid sequence of the human or chimeric IL20RB protein comprises SEQ ID NO: 113; or comprises an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 99.9% identical to SEQ ID NO: 113. In some embodiments, the amino acid sequence of the human or chimeric IL20RB protein comprises an amino acid sequence that is identical or has at least 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 99.9% identical to the amino acid sequence shown in positions 44-244, 30-254, 35-244, 35-250, 35-234, 44-254, 44-250, or 44-233 of SEQ ID NO: 113. In some embodiments, the amino acid sequence of the human or chimeric IL20RB protein comprises at least 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 99.9% of the amino acid sequence identical or identical to SEQ ID NO: 121. In some embodiments, the non-human animal is a mammal, such as a monkey or rodent (e.g., a mouse or rat). In some embodiments, the non-human animal is a mouse. In some embodiments, the endogenous IL20RB protein in the non-human animal is not expressed or is expressed at a reduced level compared to IL20RB in wild-type animals. In some embodiments, one or more cells of the non-human animal express the human or chimeric IL20RB protein.

[0719] In a fifty-fifth aspect, the present invention provides a genetically modified non-human animal or a method thereof for construction, wherein in some embodiments, the genome of the non-human animal includes a nucleotide sequence at the endogenous IL20RB locus that replaces a nucleotide sequence encoding a region of human or chimeric IL20RB with a nucleotide sequence encoding a region of human or chimeric IL20RB. In some embodiments, the nucleotide sequence encoding the region of human or chimeric IL20RB is operatively linked to an endogenous regulatory element (e.g., promoter, 5'UTR, and / or 3'UTR) of the non-human animal's endogenous IL20RB locus. In some embodiments, one or more cells of the non-human animal express human or chimeric IL20RB protein (e.g., humanized IL20RB protein). In some embodiments, the endogenous IL20RB protein of the non-human animal is not expressed or is expressed at a reduced level compared to IL20RB in wild-type animals. In some embodiments, the replaced nucleotide sequence encoding the region of endogenous IL20RB is a nucleotide sequence encoding a portion of the extracellular domain and a portion of the transmembrane domain of the endogenous IL20RB protein in the non-human animal. In some embodiments, the chimeric IL20RB protein comprises an extracellular domain, a transmembrane domain, and a cytoplasmic domain, wherein the extracellular domain is identical or identical to the extracellular domain of the human IL20RB protein by at least 50%, 60%, 70%, 80%, 90%, 95%, or 99.9%. In some embodiments, the transmembrane domain is identical or identical to the transmembrane domain of the human IL20RB protein by at least 50%, 60%, 70%, 80%, 90%, 95%, or 99.9%. In some embodiments, the cytoplasmic domain is identical or identical to the cytoplasmic domain of the non-human endogenous IL20RB protein by at least 50%, 60%, 70%, 80%, 90%, 95%, or 99.9%. In some embodiments, the extracellular domain is identical to at least 10, 20, 50, 100, 150, 190, 191, 200, 201, or 204 consecutive amino acids of the extracellular domain of the human IL20RB protein. In some embodiments, the transmembrane domain is identical to at least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or 21 consecutive amino acids of the transmembrane domain of the human IL20RB protein. Preferably, the cytoplasmic domain is identical to at least 10, 20, 50, 55, or 59 consecutive amino acids of the cytoplasmic domain of the non-human animal endogenous IL20RB protein (e.g., positions 250-308 of SEQ ID NO: 112). In some embodiments, the nucleotide sequence encoding the human or chimeric IL20RB region comprises a portion of exon 2 of the human IL20RB gene, all of exons 3 through 5, and a portion of exon 6.Preferably, the nucleotide sequence encoding the human or chimeric IL20RB region comprises at least 50-100, 100-1000, 1000-5000, and 10000-15000 bp nucleotides of exon 2, all of exons 3 to 5, and a portion of exon 6 of the human IL20RB gene, respectively. In some embodiments, the nucleotide sequence encoding the human or chimeric IL20RB region comprises portions of exons 2 to 6 of the human IL20RB gene. In some embodiments, the nucleotide sequence encoding the human or chimeric IL20RB region is identical or identical to the nucleotide sequence shown at positions 136980507-136995463 of NCBI Gene Registry Number NC_000003.12 by at least 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 99.9%. In some embodiments, the nucleotide sequence encoding the human or chimeric IL20RB region is the nucleotide sequence encoding the human or chimeric IL20RB protein. In some embodiments, the substituted nucleotide sequence encoding the endogenous IL20RB region comprises portions of exons 2 to 6 of the nonhuman animal IL20RB gene. In some embodiments, the modified IL20RB gene in the nonhuman animal genome is homozygous or heterozygous for the endogenously substituted locus.

[0720] In a fifty-sixth aspect, the present invention provides a non-human animal or a method for constructing the thereof, wherein in some embodiments, the non-human animal comprises a cell encoding at least one nucleotide sequence of a human or chimeric IL20RB protein, wherein the chimeric IL20RB protein comprises at least 50, 100, 150, 190, 191, 200, 201, 207, 210, 216, 300, 310, or 311 consecutive amino acid sequences consistent with the sequence of a human IL20RB protein. In some embodiments, the non-human animal expresses the human or chimeric IL20RB protein. The relevant limitations for the human or chimeric IL20RB protein are the same as those defined above. In some embodiments, the nucleotide sequence encoding the human or chimeric IL20RB protein is operatively linked to an endogenous IL20RB regulatory element (e.g., a promoter, 5'UTR, and / or 3'UTR). In some embodiments, the nucleotide sequence encoding the human or chimeric IL20RB protein may be integrated into the endogenous IL20RB locus of the non-human animal. In some embodiments, the human or chimeric IL20RB protein has at least one mouse IL20RB activity and / or human IL20RB activity.

[0721] In a fifty-seventh aspect, the present invention provides a genetically modified non-human animal or a method thereof, wherein in some embodiments, in at least one cell of the non-human animal, at the non-human animal endogenous IL20RB gene locus, a nucleotide sequence encoding an endogenous IL20RB region is replaced by a nucleotide sequence encoding a human or chimeric IL20RB region (or a nucleotide sequence encoding a human or chimeric IL20RB protein). In some embodiments, the endogenous IL20RB protein of the non-human animal is not expressed or is expressed at a reduced level compared to IL20RB in wild-type animals. The relevant limitations on the nucleotide sequence encoding the human or chimeric IL20RB region are the same as those described above. The relevant limitations on the human or chimeric IL20RB protein are the same as those described above. In some embodiments, the replaced nucleotide sequence encoding the endogenous IL20RB region comprises portions of exons 2 to 6 of the non-human animal IL20RB gene. In some embodiments, the nucleotide sequence encoding the human or chimeric IL20RB region is operatively linked to a non-human animal endogenous IL20RB regulatory element (e.g., a promoter, 5'UTR, and / or 3'UTR). In some embodiments, the non-human animal is a mammal, such as a monkey or a rodent (e.g., a mouse or a rat).

[0722] In a fifty-eighth aspect, the present invention provides a method for constructing a gene-modified non-human animal cell expressing human or chimeric IL20RB, the method comprising replacing, at the non-human animal endogenous IL20RB gene locus, a nucleotide sequence encoding an endogenous IL20RB region with a nucleotide sequence encoding a human or chimeric IL20RB region (or a nucleotide sequence encoding a human or chimeric IL20RB protein), thereby generating a gene-modified non-human animal cell expressing a human or chimeric IL20RB protein. The relevant limitations regarding the human or chimeric IL20RB protein are the same as those described above. The relevant limitations regarding the nucleotide sequence encoding the human or chimeric IL20RB region are the same as those described above. In some embodiments, the replaced nucleotide sequence encoding the endogenous IL20RB region is a nucleotide sequence encoding a portion of the extracellular domain and a portion of the transmembrane domain of the non-human animal endogenous IL20RB. In some embodiments, the nucleotide sequence encoding the endogenous IL20RB region comprises portions of exons 2 to 6 of the non-human animal IL20RB gene. In some embodiments, the nucleotide sequence encoding the human or chimeric IL20RB protein is operatively linked to a regulatory element of endogenous IL20RB (e.g., a promoter, 5'UTR, and / or 3'UTR). In some embodiments, the non-human animal is a mouse. In some embodiments, the non-human animal further includes nucleotide sequences of other genes encoding human or chimeric proteins selected from at least one of IL22, IL22RA1, IL10RB, IL20RA, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3, and OX40. Preferably, the IL22 is a chimeric IL22 protein; more preferably, the chimeric IL22 protein comprises any of the humanized IL22 proteins described in this invention. Preferably, the IL22RA1 is a chimeric IL22RA1 protein; more preferably, the chimeric IL22RA1 protein comprises any of the humanized IL22RA1 proteins described in this invention. Preferably, the IL10RB is a chimeric IL10RB protein; more preferably, the chimeric IL10RB protein comprises any of the humanized IL10RB proteins described in this invention. Preferably, the IL20RA is a chimeric IL20RA protein; more preferably, the chimeric IL20RA protein comprises any of the humanized IL20RA proteins described in this invention.

[0723] In a fifty-ninth aspect, the present invention provides a humanized IL20RB protein, wherein in some embodiments, the humanized IL20RB protein comprises all or part of a human IL20RB protein. In some embodiments, the humanized IL20RB protein comprises an amino acid sequence that is identical or identical in at least 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 99.9% to the amino acid sequence shown in SEQ ID NO: 113, SEQ ID NO: 113, positions 44-244, 30-254, 35-244, 35-250, 35-234, 44-254, 44-250, or 44-233. In some embodiments, the humanized IL20RB protein comprises an amino acid sequence that is identical or has at least 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 99.9% identity with the amino acid sequence shown in SEQ ID NO: 121.

[0724] In a sixtieth aspect, the present invention provides a humanized IL20RB gene, wherein in some embodiments, the humanized IL20RB gene encodes the humanized IL20RB protein. In some embodiments, the humanized IL20RB gene comprises a portion of exon 2, all of exons 3 to 5, and a portion of exon 6 of the human IL20RB gene. In some embodiments, the humanized IL20RB gene comprises portions of exons 2 to 6 of the human IL20RB gene, wherein the portion of exon 2 comprises at least a 20 bp continuous nucleotide sequence, for example, at least 20, 30, 40, 50, 60, 70, 80, 85, 86, 90, 100, 110, 120, or 127 bp continuous nucleotide sequences. The exon 6 portion contains at least 20 bp of continuous nucleotide sequence, such as at least 20, 30, 40, 50, 60, 70, 80, 85, 86, 90, 100, 110, 120, 130, 140, or 143 bp of continuous nucleotide sequence. In some embodiments, the humanized IL20RB gene comprises the entirety of exon 1, a portion of exon 2, a portion of exon 6, and the entirety of exon 7 of the non-human animal IL20RB gene. In some embodiments, the nucleotide sequence comprising the humanized IL20RB gene is identical or has at least 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 99.9% identity with the nucleotide sequence shown in SEQ ID NO: 114, 115, 116, 117, 118, 119, 120, 122, 123 or NCBI Gene Accession Number NC_000003.12 at positions 136980507-136995463.

[0725] In a sixty-first aspect, the present invention provides a targeting vector comprising: a) a 5' homologous DNA fragment (5' arm or 5' homologous arm) of the conversion region to be modified, which is selected from the genomic DNA of the IL20RB gene and has a length of 100 to 10,000 nucleotides; b) a donor region; and c) a 3' homologous DNA fragment (5' arm or 5' homologous arm) of the conversion region to be modified, which is selected from the genomic DNA of the IL20RB gene and has a length of 100 to 10,000 nucleotides.

[0726] In some embodiments, a) the DNA fragment homologous to the 5' end of the transition region to be modified is selected from a nucleotide sequence with at least 90% homology to the NCBI accession number NC_000075.7; a) the DNA fragment homologous to the 3' end of the transition region to be modified is selected from a nucleotide sequence with at least 90% homology to the NCBI accession number NC_000075.7.

[0727] In some embodiments, the length of the genomic nucleotide sequence selected for the targeting vector can exceed 0.8kb, 1kb, 2kb, 3kb, 4kb, 5kb, 6kb, 7kb, 8kb, 9kb, 10kb, 15kb, 16kb, 17kb, 18kb, 19kb, or 20kb.

[0728] In some embodiments, the conversion region to be altered is located on exons 1 to 7 of the non-human animal endogenous IL20RB gene, preferably on exons 2 to 6.

[0729] In some embodiments, the 5' homologous arm comprises SEQ ID NO: 114 and / or 122.

[0730] In some embodiments, the 3' homologous arm comprises SEQ ID NO: 115 and / or 123.

[0731] In some embodiments, the donor sequence comprises a human sequence, such as bits 136980507-136995463 of NCBI Gene Accession Number NC_000003.12.

[0732] In some embodiments, the targeting vector comprises one or more marker genes (or resistance genes), such as resistance genes for positive clone selection or genes encoding negative selection markers. In some embodiments, the resistance gene for positive clone selection is the neomycin phosphotransferase coding sequence Neo. Preferably, the targeting vector further comprises two co-directed Frt recombination sites flanking the marker gene. In some embodiments, the gene encoding the negative selection marker comprises the gene encoding the diphtheria toxin A subunit (DTA).

[0733] This invention also provides vectors for constructing humanized animal models or knockout models. In some embodiments, the vector contains an sgRNA sequence that targets the IL20RB gene. In some embodiments, the target site of the sgRNA in the conversion region to be altered is unique and satisfies the sequence arrangement rule of 5'-NNN(20)-NGG3' or 5'-CCN-N(20)-3'. In some embodiments, the target site of the sgRNA targeting the non-human IL20RB gene is located on exon 2 and / or exon 6 of the IL20RB gene.

[0734] Preferably, the target site sequence of the sgRNA targeting the IL20RB gene in non-human animals includes SEQ ID NO: 124 and / or 125.

[0735] Preferably, the sgRNA targeting the non-human animal IL20RB gene includes nucleotide sequences as shown in SEQ ID NO: 126 and 128, or nucleotide sequences as shown in SEQ ID NO: 127 and 129, or nucleotide sequences as shown in SEQ ID NO: 130 and 132, or nucleotide sequences as shown in SEQ ID NO: 131 and 133.

[0736] In some embodiments, the present invention relates to plasmid constructs containing sgRNA (sgRNA vectors, such as pT7-sgRNA vectors) and / or cells containing said constructs.

[0737] The present invention also relates to cells comprising the targeting vector or sgRNA vector as described above.

[0738] In some embodiments, the present invention also provides a non-human mammalian cell having any of the above-described targeting vectors and one or more in vitro transcripts from the plasmid construction system described in this application. In some embodiments, the non-human mammalian cell contains Cas9 mRNA or its in vitro transcript.

[0739] In some embodiments, the genes in the non-human mammalian cells are heterozygous. In some embodiments, the genes in the non-human mammalian cells are homozygous.

[0740] In some embodiments, the non-human mammalian cell is a mouse cell. In some embodiments, the non-human mammalian cell is a fertilized egg cell. In some embodiments, the non-human mammalian cell is an embryonic stem cell. In some embodiments, the non-human mammalian cell is any type of cell capable of expressing IL20RB.

[0741] According to a sixty-second aspect of the present invention, a nucleic acid comprising any one of the following:

[0742] 1) A nucleotide sequence encoding the modified IL22RA1 protein; preferably, the nucleic acid further comprises SEQ ID NO: 30; preferably, the mRNA transcribed from the nucleic acid comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 31; B) the identity with the nucleotide sequence shown in SEQ ID NO: 31 is at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99%; C) the difference from the nucleotide sequence shown in SEQ ID NO: 31 is no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or, D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 31, including substitutions, deletions, and / or insertions of one or more nucleotides;

[0743] 2) A nucleotide sequence encoding the human or modified IL22 protein; preferably, the nucleic acid further comprises SEQ ID NO: 5; preferably, the mRNA transcribed from the nucleic acid comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 6; B) at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 6; C) differing from the nucleotide sequence shown in SEQ ID NO: 6 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 6, including substitutions, deletions, and / or insertions of one or more nucleotides;

[0744] 3) A nucleotide sequence encoding the modified IL20RA protein; preferably, the nucleic acid further comprises the nucleotide sequence shown in positions 137004735-137017092 of NCBI Gene Accession Number NC_000006.12; preferably, the mRNA transcribed from the nucleic acid comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 90; B) the identity with the nucleotide sequence shown in SEQ ID NO: 90 is at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99%; C) the difference from the nucleotide sequence shown in SEQ ID NO: 90 is no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or, D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 90, including substitutions, deletions, and / or insertions of one or more nucleotides;

[0745] 4) A nucleotide sequence encoding the modified IL20RB protein; preferably, the nucleic acid further comprises the nucleotide sequence shown in positions 136980507-136995463 of NCBI Gene Accession Number NC_000003.12; preferably, the mRNA transcribed from the nucleic acid comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 120; B) the identity with the nucleotide sequence shown in SEQ ID NO: 120 is at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99%; C) the difference from the nucleotide sequence shown in SEQ ID NO: 120 is no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 120, including substitutions, deletions, and / or insertions of one or more nucleotides;

[0746] 5) The nucleotide sequence encoding the modified IL10RB protein; preferably, the nucleic acid further comprises the nucleotide sequence or a variant thereof shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9; wherein the variant comprises one or more of the following: C>T mutation at position 33276503, G>A mutation at position 33276818, or TTTGTGG>TTG mutation at positions 33277048-33277053; preferably, the mRNA transcribed from the nucleic acid comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 61; B) the identity with the nucleotide sequence shown in SEQ ID NO: 61 is at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99%; C) the identity with the nucleotide sequence shown in SEQ ID NO: 61. The nucleotide sequence shown in SEQ ID NO: 61 differs by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2 or no more than 1 nucleotide; or, D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 61, including substitutions, deletions and / or insertions of one or more nucleotides.

[0747] According to a sixty-third aspect of the present invention, a polypeptide is provided, said polypeptide comprising any one of the following:

[0748] 1) Encoded by the genetically modified endogenous IL22RA1 gene;

[0749] 2) Encoded by the genetically modified endogenous IL22 gene;

[0750] 3) Encoded by the genetically modified endogenous IL20RA gene;

[0751] 4) Encoded by the genetically modified endogenous IL20RB gene;

[0752] 5) Encoded by the genetically modified endogenous IL10RB gene;

[0753] Preferably, the polypeptide is encoded by the aforementioned nucleic acid.

[0754] In its sixty-fourth aspect, the present invention provides a non-human animal genome modified with dual genes IL22 and IL22RA1.

[0755] The non-human animal genome includes the genetically modified endogenous IL22 gene encoding the human or modified IL22 protein, and the genetically modified endogenous IL22RA1 gene encoding the modified IL22RA1 protein.

[0756] Preferably, the non-human animal expresses the human or modified IL22 protein and the modified IL22RA1 protein.

[0757] Preferably, the non-human animal comprises the aforementioned non-human animal genome.

[0758] Preferably, the non-human animal genome contains a wild-type endogenous IL10RB gene encoding the wild-type endogenous IL10RB protein.

[0759] In one specific embodiment of the present invention, the non-human animal genome includes:

[0760] I) The wild-type endogenous IL10RB gene encoding the wild-type endogenous IL10RB protein; and,

[0761] II) Genetically modified endogenous IL22 genes encoding human or modified IL22 proteins and genetically modified endogenous IL22RA1 genes encoding modified IL22RA1 proteins.

[0762] Preferably, in non-human animals containing the non-human animal genome, the differentiation, development, and distribution of leukocyte subtypes and / or T cell subtypes are not significantly different from those in non-human animals expressing wild-type endogenous IL22 and wild-type endogenous IL22RA1.

[0763] Among them, white blood cell subtypes include one or more of B cells, T cells, NK cells, dendritic cells (DCs), granulocytes, macrophages, or monocytes. T cell subtypes include one or more of CD4+ T cells, CD8+ T cells, or Tregs.

[0764] Preferably, the leukocytes or T cells include leukocytes or T cells derived from the spleen, blood, or lymph nodes.

[0765] Preferably, in non-human animals containing the non-human animal genome, the expression levels of CXCL1 and IL6 are not significantly different from those in non-human animals expressing wild-type endogenous IL22, wild-type endogenous IL10RB, and wild-type endogenous IL22RA1.

[0766] In a sixty-fifth aspect of the present invention, a non-human animal with dual IL22 and IL22RA1 gene modification is provided, said non-human animal comprising the non-human animal genome of the sixty-fourth aspect.

[0767] In a sixty-sixth aspect of the present invention, a non-human animal genome modified with dual genes of IL22RA1 and IL10RB is provided, the non-human animal genome comprising a genetically modified endogenous IL22RA1 gene encoding the modified IL22RA1 protein and a genetically modified endogenous IL10RB gene encoding the modified IL10RB protein.

[0768] Preferably, the non-human animal expresses the modified IL22RA1 protein and the modified IL10RB protein.

[0769] Preferably, the non-human animal genome contains a wild-type endogenous IL22 gene encoding the wild-type endogenous IL22 protein.

[0770] In one specific embodiment of the present invention, the non-human animal genome includes:

[0771] I) The wild-type endogenous IL22 gene encoding the wild-type endogenous IL22 protein; and,

[0772] II) Genetically modified endogenous IL10RB gene encoding modified IL10RB protein and genetically modified endogenous IL22RA1 gene encoding modified IL22RA1 protein.

[0773] Preferably, in non-human animals containing the non-human animal genome, the differentiation, development, and distribution of leukocyte subtypes and / or T cell subtypes are not significantly different from those in non-human animals expressing wild-type endogenous IL10RB and wild-type endogenous IL22RA1.

[0774] Among them, white blood cell subtypes include one or more of B cells, T cells, NK cells, dendritic cells (DCs), granulocytes, macrophages, or monocytes. T cell subtypes include one or more of CD4+ T cells, CD8+ T cells, or Tregs.

[0775] Preferably, the leukocytes or T cells include leukocytes or T cells derived from the spleen, blood, or lymph nodes.

[0776] Preferably, in non-human animals containing the non-human animal genome, the expression levels of CXCL1 and IL6 are not significantly different from those in non-human animals expressing wild-type endogenous IL22, wild-type endogenous IL10RB, and wild-type endogenous IL22RA1.

[0777] In a sixty-seventh aspect of the present invention, a non-human animal with dual gene modification of IL22RA1 and IL10RB is provided, wherein the non-human animal comprises the non-human animal genome described in the sixty-sixth aspect.

[0778] In its sixty-eighth aspect, the present invention provides a non-human animal genome modified with three genes: IL22, IL22RA1, and IL10RB.

[0779] The non-human animal genome contains the aforementioned genetically modified endogenous IL22 gene, genetically modified endogenous IL22RA1 gene, and genetically modified endogenous IL10RB gene.

[0780] Preferably, in non-human animals containing the non-human animal genome, the expression levels of CXCL1 and IL6 are not significantly different from those in non-human animals expressing wild-type endogenous IL22, wild-type endogenous IL10RB, and wild-type endogenous IL22RA1.

[0781] The present invention also provides a non-human animal comprising the non-human animal genome described in aspect sixty-eight.

[0782] In its sixty-ninth aspect, the present invention provides a non-human animal genome modified with three genes: IL10RB, IL10, and IL10RA.

[0783] The non-human animal genome contains the aforementioned genetically modified endogenous IL10RB gene, genetically modified endogenous IL10 gene, and genetically modified endogenous IL10RA gene.

[0784] Preferably, in non-human animals containing the non-human animal genome, the expression levels of CXCL1 and IL6 are not significantly different from those in non-human animals expressing wild-type endogenous IL10, wild-type endogenous IL10RB, and wild-type endogenous IL10RA1.

[0785] The present invention also provides a non-human animal comprising the non-human animal genome described in aspect sixty-nine.

[0786] In a seventieth aspect of the present invention, a non-human animal genome modified with five genes, namely IL22, IL22RA1, IL10RB, IL10, and IL10RA, is provided.

[0787] The non-human animal genome contains the aforementioned genetically modified endogenous IL22 gene, genetically modified endogenous IL22RA1 gene, genetically modified endogenous IL10RB gene, genetically modified endogenous IL10 gene, and genetically modified endogenous IL10RA gene.

[0788] Preferably, in non-human animals containing the non-human animal genome, the expression levels of CXCL1 and IL6 are not significantly different from those in non-human animals expressing wild-type endogenous IL10RB, wild-type endogenous IL22RA1, wild-type endogenous IL22, wild-type endogenous IL10 and wild-type endogenous IL10RA.

[0789] In its seventy-first aspect, the present invention provides a non-human animal comprising the non-human animal genome described in the seventieth aspect.

[0790] In a seventy-second aspect, the present invention provides a method for constructing a non-human animal modified with two or more of the genes IL22, IL22RA1, IL10RB, IL10, IL10RA, IL20RA, or IL20RB, wherein the modification includes gene humanization, preferably, the non-human animal expressing two or more of the following: humanized IL22 protein, humanized IL22RA1 protein, humanized IL10RB protein, humanized IL20RA protein, modified IL10 protein, modified IL10RA protein, or humanized IL20RB protein.

[0791] Preferably, the construction method includes:

[0792] To mate or in-vitro fertilize non-human animals;

[0793] Alternatively, gene editing can be performed on non-human animals to obtain non-human animals with multiple gene modifications.

[0794] Preferably, the humanized IL22 protein is selected from the above-mentioned humanized IL22 protein, the humanized IL22RA1 protein is selected from the above-mentioned humanized IL22RA1 protein, the humanized IL10RB protein is selected from the above-mentioned humanized IL10RB protein, the humanized IL20RA protein is selected from the above-mentioned humanized IL20RA protein, and the humanized IL20RB protein is selected from the above-mentioned humanized IL20RB protein.

[0795] In its seventy-third aspect, the present invention provides a method for constructing a multi-gene modified non-human animal, comprising the following steps:

[0796] I) Provide the above-mentioned non-human animals, or non-human animals obtained using the above-mentioned construction method;

[0797] II) The non-human animals provided in step I) are mated with other genetically modified non-human animals, fertilized in vitro, or directly genetically edited, and then screened to obtain multi-gene-modified non-human animals.

[0798] Preferably, the other genetically modified non-human animals include non-human animals modified with genes IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3, or OX40.

[0799] Preferably, the multi-gene modified non-human animal is a two-gene humanized non-human animal, a three-gene humanized non-human animal, a four-gene humanized non-human animal, a five-gene humanized non-human animal, a six-gene humanized non-human animal, or a seven-gene humanized non-human animal.

[0800] Preferably, in the genome of the multi-gene modified non-human animal, each of the multiple humanized genes can be homozygous or heterozygous for the modification of the endogenous locus.

[0801] In its seventy-fourth aspect, the present invention provides a cell, tissue, or organ expressing the aforementioned humanized IL22 protein, the aforementioned humanized IL22RA1 protein, the aforementioned humanized IL10RB protein, the aforementioned humanized IL20RA protein, the aforementioned humanized IL20RB protein, or any of the aforementioned polypeptides. Alternatively, the genome of the cell, tissue, or organ contains the aforementioned humanized IL22 gene, the aforementioned humanized IL22RA1 gene, the aforementioned humanized IL1...

Claims

A non-human animal genome, characterized in that, The non-human animal or the non-human animal genome contains a genetically modified endogenous IL22RA1 gene encoding a modified IL22RA1 protein, and the non-human animal expresses the modified IL22RA1 protein. The non-human animal or non-human animal genome according to claim 1 is characterized in that, The non-human animal or non-human animal genome contains a nucleotide sequence that replaces the corresponding segment of the non-human animal endogenous IL22RA1 with a nucleotide sequence that encodes the human IL22RA1 segment. The non-human animal or non-human animal genome according to claim 1 or 2 is characterized in that, The modified IL22RA1 protein comprises a cytoplasmic domain, a transmembrane domain, and an extracellular domain. Preferably, the modified IL22RA1 protein contains an extracellular domain that is substantially the same as that of the human IL22RA1 protein. The non-human animal or non-human animal genome according to any one of claims 1-3 is characterized in that, The extracellular domain may comprise all or part of the amino acid sequence shown in positions 16-228 or 17-216 of SEQ ID NO: 27; or, may comprise an amino acid sequence with an identity of at least 77%, 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown in positions 16-228 or 17-216 of SEQ ID NO: 27; or, may comprise an amino acid sequence differing from the amino acid sequence shown in positions 16-228 or 17-216 of SEQ ID NO: 27 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, may comprise an amino acid sequence with an identity of positions 16-228 or 17-216 of SEQ ID NO: 27, including substitutions, deletions, and / or insertions of one or more amino acid residues. The non-human animal or non-human animal genome according to any one of claims 1-4 is characterized in that, The non-human animal is a rodent, more preferably a rat or mouse. The non-human animal or non-human animal genome according to any one of claims 1-5 is characterized in that, The modified IL22RA1 protein contains essentially the same cytoplasmic domains as the endogenous IL22RA1 protein in non-human animals. Preferably, the amino acid sequence of the cytoplasmic domain comprises the amino acid sequence shown in positions 252-581 of SEQ ID NO: 26; or, comprises an amino acid sequence with at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in positions 252-581 of SEQ ID NO: 26; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 252-581 of SEQ ID NO: 26 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with substitutions, deletions, and / or insertions of one or more amino acid residues as shown in positions 252-581 of SEQ ID NO:

26. The non-human animal or non-human animal genome according to any one of claims 1-6 is characterized in that, The modified IL22RA1 protein contains a transmembrane domain that is substantially the same as that of the non-human endogenous IL22RA1 protein. Preferably, the amino acid sequence of the transmembrane domain comprises the amino acid sequence shown in positions 231-251 of SEQ ID NO: 26; or, comprises an amino acid sequence with an identity of at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown in positions 231-251 of SEQ ID NO: 26; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 231-251 of SEQ ID NO: 26 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with an identity of positions 231-251 of SEQ ID NO: 26, including substitutions, deletions, and / or insertions of one or more amino acid residues. The non-human animal or non-human animal genome according to any one of claims 1-7 is characterized in that, The modified IL22RA1 protein has an amino acid sequence comprising the amino acid sequence shown in SEQ ID NO: 32; or, comprising an amino acid sequence with at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in SEQ ID NO: 32; or, comprising an amino acid sequence differing from the amino acid sequence shown in SEQ ID NO: 32 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprising an amino acid sequence similar to that shown in SEQ ID NO: 32, including substitutions, deletions, and / or insertions of one or more amino acid residues. The non-human animal or non-human animal genome according to any one of claims 1-8 is characterized in that, The nucleotide sequence encoding the human IL22RA1 region contains a portion of exon 2, all of exon 3, all of exon 4, and a portion of exon 5 of the human IL22RA1 gene. The non-human animal or non-human animal genome according to any one of claims 1-9 is characterized in that, The nucleotide sequence encoding the human IL22RA1 region comprises the nucleotide sequence shown in SEQ ID NO: 30; or, comprises a nucleotide sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 30; or, comprises a nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 30 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or, comprises a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 30, including substitutions, deletions, and / or insertions of one or more nucleotides. The non-human animal or non-human animal genome according to any one of claims 1-10 is characterized in that, The mRNA transcribed from the nucleotide sequence encoding the modified IL22RA1 protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 31; B) the nucleotide sequence being at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identical to the nucleotide sequence shown in SEQ ID NO: 31; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 31 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 31, including substitutions, deletions, and / or insertions of one or more nucleotides. The non-human animal or non-human animal genome according to any one of claims 1-11 is characterized in that, The expression of the modified IL22RA1 protein is controlled by endogenous IL22RA1 regulatory elements (e.g., promoter, 5'UTR and / or 3'UTR). The non-human animal or non-human animal genome according to any one of claims 1-12 is characterized in that, The non-human animal or non-human animal genome is homozygous or heterozygous for the genetically modified endogenous IL22RA1 gene. The non-human animal or non-human animal genome according to any one of claims 1-13 is characterized in that, The non-human animal or non-human animal genome contains endogenous genes encoding human or modified other genes, preferably, the other genes are selected from at least one of IL22, IL10RB, IL20RA, IL20RB, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3 and OX40. A method for preparing non-human animals, characterized in that, The non-human animal contains a genetically modified endogenous IL22RA1 gene encoding a modified IL22RA1 protein, and the non-human animal expresses the modified IL22RA1 protein. The method according to claim 15, characterized in that, The non-human animal or the genome of the non-human animal contains a nucleotide sequence that replaces the nucleotide sequence encoding the corresponding segment of the non-human animal's endogenous IL22RA1 with a nucleotide sequence encoding the human IL22RA1 segment. The method according to claim 15 or 16 is characterized in that, The modified IL22RA1 protein comprises a cytoplasmic domain, a transmembrane domain, and an extracellular domain. Preferably, the modified IL22RA1 protein contains an extracellular domain that is substantially the same as that of the human IL22RA1 protein; Preferably, all or part of the extracellular domain comprises the amino acid sequence shown at positions 16-228 or 17-216 of SEQ ID NO: 27; or, comprises an amino acid sequence with an identity of at least 77%, 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown at positions 16-228 or 17-216 of SEQ ID NO: 27; or, comprises an amino acid sequence differing from the amino acid sequence shown at positions 16-228 or 17-216 of SEQ ID NO: 27 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with an identity of positions 16-228 or 17-216 of SEQ ID NO: 27, including substitutions, deletions, and / or insertions of one or more amino acid residues. The method according to any one of claims 15-17 is characterized in that, The non-human animal is a rodent, more preferably a rat or mouse. The method according to any one of claims 15-18, characterized in that, The modified IL22RA1 protein contains essentially the same cytoplasmic domains as the endogenous IL22RA1 protein in non-human animals. Preferably, the amino acid sequence of the cytoplasmic domain comprises the amino acid sequence shown in positions 252-581 of SEQ ID NO: 26; or, comprises an amino acid sequence with at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in positions 252-581 of SEQ ID NO: 26; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 252-581 of SEQ ID NO: 26 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with substitutions, deletions, and / or insertions of one or more amino acid residues as shown in positions 252-581 of SEQ ID NO:

26. The method according to any one of claims 15-19, characterized in that, The modified IL22RA1 protein contains a transmembrane domain that is substantially the same as that of the non-human endogenous IL22RA1 protein. Preferably, the amino acid sequence of the transmembrane domain comprises the amino acid sequence shown in positions 231-251 of SEQ ID NO: 26; or, comprises an amino acid sequence with an identity of at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown in positions 231-251 of SEQ ID NO: 26; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 231-251 of SEQ ID NO: 26 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with an identity of positions 231-251 of SEQ ID NO: 26, including substitutions, deletions, and / or insertions of one or more amino acid residues. The method according to any one of claims 15-20 is characterized in that, The modified IL22RA1 protein has an amino acid sequence comprising the amino acid sequence shown in SEQ ID NO: 32; or, comprising an amino acid sequence with at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in SEQ ID NO: 32; or, comprising an amino acid sequence differing from the amino acid sequence shown in SEQ ID NO: 32 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprising an amino acid sequence similar to that shown in SEQ ID NO: 32, including substitutions, deletions, and / or insertions of one or more amino acid residues. The method according to any one of claims 15-21 is characterized in that, The nucleotide sequence encoding the human IL22RA1 region contains a portion of exon 2, all of exon 3, all of exon 4, and a portion of exon 5 of the human IL22RA1 gene. The method according to any one of claims 15-22 is characterized in that, The nucleotide sequence encoding the human IL22RA1 region comprises the nucleotide sequence shown in SEQ ID NO: 30; or, comprises a nucleotide sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 30; or, comprises a nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 30 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or, comprises a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 30, including substitutions, deletions, and / or insertions of one or more nucleotides. The method according to any one of claims 15-23 is characterized in that, The mRNA transcribed from the nucleotide sequence encoding the modified IL22RA1 protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 31; B) the nucleotide sequence being at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identical to the nucleotide sequence shown in SEQ ID NO: 31; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 31 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 31, including substitutions, deletions, and / or insertions of one or more nucleotides. The method according to any one of claims 15-24 is characterized in that, The expression of the modified IL22RA1 protein is controlled by endogenous IL22RA1 regulatory elements (e.g., promoter, 5'UTR and / or 3'UTR). The method according to any one of claims 15-25 is characterized in that, The genome of the non-human animal is homozygous or heterozygous for the genetically modified endogenous IL22RA1 gene. The method according to any one of claims 15-26 is characterized in that, The non-human animal or the genome of the non-human animal contains an endogenous gene encoding a human or modified other gene, preferably, the other gene is selected from at least one of IL22, IL10RB, IL20RA, IL20RB, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3 and OX40. A non-human animal genome, characterized in that, The non-human animal or non-human animal genome contains a genetically modified endogenous IL22 gene encoding a human or modified IL22 protein, and the non-human animal expresses the human or modified IL22 protein. The non-human animal or non-human animal genome according to claim 28 is characterized in that, The non-human animal or non-human animal genome contains a nucleotide sequence that replaces the corresponding nucleotide sequence encoding the non-human animal endogenous IL22 with a nucleotide sequence encoding the human IL22 region. The non-human animal or non-human animal genome according to any one of claims 28-29 is characterized in that, The human or modified IL22 protein comprises the amino acid sequence shown in SEQ ID NO: 2; or, comprises an amino acid sequence that is at least 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identical to the amino acid sequence shown in SEQ ID NO: 2; or, comprises an amino acid sequence that differs from the amino acid sequence shown in SEQ ID NO: 2 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence that is identical to the amino acid sequence shown in SEQ ID NO: 2, including substitutions, deletions, and / or insertions of one or more amino acid residues. The non-human animal or non-human animal genome according to any one of claims 28-30 is characterized in that, The nucleotide sequence encoding the human IL22 region includes a portion of exon 2, the entirety of exon 3, the entirety of exon 4, the entirety of exon 5, and a portion of exon 6 of the human IL22 gene. The non-human animal or non-human animal genome according to any one of claims 28-31 is characterized in that, The nucleotide sequence encoding the human IL22 region comprises the nucleotide sequence shown in SEQ ID NO: 5; or, comprises a nucleotide sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 5; or, comprises a nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 5 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or, comprises a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 5, including substitutions, deletions, and / or insertions of one or more nucleotides. The non-human animal or non-human animal genome according to any one of claims 28-32 is characterized in that, The mRNA transcribed from the nucleotide sequence encoding the human or modified IL22 protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 6; B) the nucleotide sequence being at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identical to the nucleotide sequence shown in SEQ ID NO: 6; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 6 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 6, including substitutions, deletions, and / or insertions of one or more nucleotides. The non-human animal or non-human animal genome according to any one of claims 28-33 is characterized in that, The expression of human or modified IL22 proteins is controlled by endogenous IL22 regulatory elements (e.g., promoters, 5'UTR, and / or 3'UTR). The non-human animal or non-human animal genome according to any one of claims 28-34 is characterized in that, The non-human animal or non-human animal genome is homozygous or heterozygous for the genetically modified endogenous IL22 gene. The non-human animal or non-human animal genome according to any one of claims 28-35 is characterized in that, The non-human animal mentioned is a rodent, preferably a rat or mouse. The non-human animal or non-human animal genome according to any one of claims 28-36 is characterized in that, The non-human animal or non-human animal genome contains endogenous genes encoding human or modified other genes, preferably, the other genes are selected from at least one of IL22RA1, IL10RB, IL20RA, IL20RB, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3 and OX40. A method for preparing non-human animals, characterized in that, The non-human animal contains a genetically modified endogenous IL22 gene encoding a human or modified IL22 protein, and the non-human animal expresses the human or modified IL22 protein. The method according to claim 38, characterized in that, The non-human animal or the genome of the non-human animal contains a nucleotide sequence that replaces the nucleotide sequence encoding the corresponding segment of endogenous IL22 in the non-human animal with a nucleotide sequence encoding the human IL22 segment. The method according to any one of claims 38-39 is characterized in that, The human or modified IL22 protein comprises the amino acid sequence shown in SEQ ID NO: 2; or, comprises an amino acid sequence that is at least 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identical to the amino acid sequence shown in SEQ ID NO: 2; or, comprises an amino acid sequence that differs from the amino acid sequence shown in SEQ ID NO: 2 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence that is identical to the amino acid sequence shown in SEQ ID NO: 2, including substitutions, deletions, and / or insertions of one or more amino acid residues. The method according to any one of claims 38-40, characterized in that, The nucleotide sequence encoding the human IL22 region includes a portion of exon 2, the entirety of exon 3, the entirety of exon 4, the entirety of exon 5, and a portion of exon 6 of the human IL22 gene. The method according to any one of claims 38-41, characterized in that, The nucleotide sequence encoding the human IL22 region comprises the nucleotide sequence shown in SEQ ID NO: 5; or, comprises a nucleotide sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 5; or, comprises a nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 5 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or, comprises a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 5, including substitutions, deletions, and / or insertions of one or more nucleotides. The method according to any one of claims 38-42, characterized in that, The mRNA transcribed from the nucleotide sequence encoding the human or modified IL22 protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 6; B) the nucleotide sequence being at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identical to the nucleotide sequence shown in SEQ ID NO: 6; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 6 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 6, including substitutions, deletions, and / or insertions of one or more nucleotides. The method according to any one of claims 38-43 is characterized in that, The expression of human or modified IL22 proteins is controlled by endogenous IL22 regulatory elements (e.g., promoters, 5'UTR, and / or 3'UTR). The method according to any one of claims 38-44, characterized in that, The non-human animal or non-human animal genome is homozygous or heterozygous for the genetically modified endogenous IL22 gene. The method according to any one of claims 38-45, characterized in that, The non-human animal mentioned is a rodent, preferably a rat or mouse. The method according to any one of claims 38-46, characterized in that, The non-human animal or the genome of the non-human animal contains an endogenous gene encoding a human or modified other gene, preferably, the other gene is selected from at least one of IL22RA1, IL10RB, IL20RA, IL20RB, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3 and OX40. A non-human animal genome, characterized in that, The non-human animal or the non-human animal genome contains a genetically modified endogenous IL20RA gene encoding a modified IL20RA protein, and the non-human animal expresses the modified IL20RA protein. The non-human animal or non-human animal genome according to claim 48 is characterized in that, The non-human animal or non-human animal genome contains a nucleotide sequence that replaces the corresponding region encoding the non-human animal endogenous IL20RA with a nucleotide sequence encoding the human IL20RA region. The non-human animal or non-human animal genome according to any one of claims 48-49 is characterized in that, The modified IL20RA protein comprises a cytoplasmic domain, a transmembrane domain, and an extracellular domain; Preferably, the modified IL20RA protein contains an extracellular domain substantially the same as that of the human IL20RA protein; Preferably, all or part of the extracellular domain comprises the amino acid sequence shown in positions 30-250, 34-250, 30-247, or 34-247 of SEQ ID NO: 83; or, comprises an amino acid sequence with an identity of at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with ...10, 9, 8, 7, 6, 5, 4, 3, 2, or at least 1 amino acid with the amino acid sequence shown in positions 30-250, 34-250, 30-247, or 34-247 of SEQ ID NO: 83; or, comprises an amino acid sequence with an identity of at least 10, 9, 8, 7, 6, 5, 4, 3, 2, or at least 1 amino acid with the amino acid sequence shown in positions 30-250, 34-250, 30-247, or 34-247 of SEQ ID NO: 83; or, comprises an amino acid sequence with an identity of at least 10, 9, 8, 7, NO:83 indicates an amino acid sequence including substitutions, deletions, and / or insertions of one or more amino acid residues, as shown in positions 30-250, 34-250, 30-247, or 34-247. The non-human animal or non-human animal genome according to any one of claims 48-50 is characterized in that, The non-human animal mentioned is a rodent, preferably a rat or mouse. The non-human animal or non-human animal genome according to any one of claims 48-51 is characterized in that, The modified IL20RA protein contains essentially the same cytoplasmic domain as the endogenous IL20RA protein in non-human animals. Preferably, the amino acid sequence of the cytoplasmic domain comprises the amino acid sequence shown in positions 275-546 of SEQ ID NO: 82; or, comprises an amino acid sequence with at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in positions 275-546 of SEQ ID NO: 82; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 275-546 of SEQ ID NO: 82 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with positions 275-546 of SEQ ID NO: 82, including substitutions, deletions, and / or insertions of one or more amino acid residues. The non-human animal or non-human animal genome according to any one of claims 48-52 is characterized in that, The modified IL20RA protein contains a transmembrane domain that is substantially the same as that of the non-human endogenous IL20RA protein. Preferably, the amino acid sequence of the transmembrane domain comprises the amino acid sequence shown in positions 254-274 of SEQ ID NO: 82; or, comprises an amino acid sequence with at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in positions 254-274 of SEQ ID NO: 82; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 254-274 of SEQ ID NO: 82 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with positions 254-274 of SEQ ID NO: 82, including substitutions, deletions, and / or insertions of one or more amino acid residues. The non-human animal or non-human animal genome according to any one of claims 48-53 is characterized in that, The modified IL20RA protein has an amino acid sequence comprising the amino acid sequence shown in SEQ ID NO: 91; or, comprising an amino acid sequence with at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in SEQ ID NO: 91; or, comprising an amino acid sequence differing from the amino acid sequence shown in SEQ ID NO: 91 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprising an amino acid sequence similar to that shown in SEQ ID NO: 91, including substitutions, deletions, and / or insertions of one or more amino acid residues. The non-human animal or non-human animal genome according to any one of claims 48-54 is characterized in that, The nucleotide sequence encoding the human IL20RA region includes a portion of exon 2, the entirety of exon 3, the entirety of exon 4, the entirety of exon 5, and a portion of exon 6 of the human IL20RA gene. The non-human animal or non-human animal genome according to any one of claims 48-55 is characterized in that, The nucleotide sequence encoding the human IL20RA region comprises the nucleotide sequence shown in positions 137004735-137017092 of NCBI Gene Accession Number NC_000006.12; or, comprises a nucleotide sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, or 98% identity to the nucleotide sequence shown in positions 137004735-137017092 of NCBI Gene Accession Number NC_000006.

12. Or at least 99% of the nucleotide sequence; or, a nucleotide sequence that differs from the nucleotide sequence shown in positions 137004735-137017092 of NCBI Gene Accession Number NC_000006.12 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2 or no more than 1 nucleotide; or, a nucleotide sequence that includes substitutions, deletions and / or insertions of one or more nucleotides as shown in the nucleotide sequence shown in positions 137004735-137017092 of NCBI Gene Accession Number NC_000006.

12. The non-human animal or non-human animal genome according to any one of claims 48-56 is characterized in that, The mRNA transcribed from the nucleotide sequence encoding the modified IL20RA protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 90; B) the nucleotide sequence being at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identical to the nucleotide sequence shown in SEQ ID NO: 90; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 90 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 90, including substitutions, deletions, and / or insertions of one or more nucleotides. The non-human animal or non-human animal genome according to any one of claims 48-57 is characterized in that, The expression of the modified IL20RA protein is controlled by endogenous IL20RA regulatory elements (e.g., promoter, 5'UTR and / or 3'UTR). The non-human animal or non-human animal genome according to any one of claims 48-58 is characterized in that, The non-human animal or non-human animal genome is homozygous or heterozygous for the genetically modified endogenous IL20RA gene. The non-human animal or non-human animal genome according to any one of claims 48-59 is characterized in that, The non-human animal or non-human animal genome contains endogenous genes encoding human or modified other genes, preferably, the other genes are selected from at least one of IL22, IL22RA1, IL10RB, IL20RB, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3 and OX40. A method for preparing non-human animals, characterized in that, The non-human animal contains a genetically modified endogenous IL20RA gene encoding a modified IL20RA protein, and the non-human animal expresses the modified IL20RA protein. The method according to claim 61, characterized in that, The non-human animal or the genome of the non-human animal contains a nucleotide sequence that replaces the corresponding region encoding the non-human animal's endogenous IL20RA with a nucleotide sequence encoding the human IL20RA region. The method according to any one of claims 61-62 is characterized in that, The modified IL20RA protein comprises a cytoplasmic domain, a transmembrane domain, and an extracellular domain; Preferably, the modified IL20RA protein contains an extracellular domain substantially the same as that of the human IL20RA protein; Preferably, all or part of the extracellular domain comprises the amino acid sequence shown in positions 30-250, 34-250, 30-247, or 34-247 of SEQ ID NO: 83; or, comprises an amino acid sequence with an identity of at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with ...10, 9, 8, 7, 6, 5, 4, 3, 2, or at least 1 amino acid with the amino acid sequence shown in positions 30-250, 34-250, 30-247, or 34-247 of SEQ ID NO: 83; or, comprises an amino acid sequence with an identity of at least 10, 9, 8, 7, 6, 5, 4, 3, 2, or at least 1 amino acid with the amino acid sequence shown in positions 30-250, 34-250, 30-247, or 34-247 of SEQ ID NO: 83; or, comprises an amino acid sequence with an identity of at least 10, 9, 8, 7, NO:83 indicates an amino acid sequence including substitutions, deletions, and / or insertions of one or more amino acid residues, as shown in positions 30-250, 34-250, 30-247, or 34-247. The method according to any one of claims 61-63 is characterized in that, The non-human animal mentioned is a rodent, preferably a rat or mouse. The method according to any one of claims 61-64, characterized in that, The modified IL20RA protein contains essentially the same cytoplasmic domain as the endogenous IL20RA protein in non-human animals. Preferably, the amino acid sequence of the cytoplasmic domain comprises the amino acid sequence shown in positions 275-546 of SEQ ID NO: 82; or, comprises an amino acid sequence with at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in positions 275-546 of SEQ ID NO: 82; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 275-546 of SEQ ID NO: 82 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with positions 275-546 of SEQ ID NO: 82, including substitutions, deletions, and / or insertions of one or more amino acid residues. The method according to any one of claims 61-65, characterized in that, The modified IL20RA protein contains a transmembrane domain that is substantially the same as that of the non-human endogenous IL20RA protein. Preferably, the amino acid sequence of the transmembrane domain comprises the amino acid sequence shown in positions 254-274 of SEQ ID NO: 82; or, comprises an amino acid sequence with at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in positions 254-274 of SEQ ID NO: 82; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 254-274 of SEQ ID NO: 82 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with positions 254-274 of SEQ ID NO: 82, including substitutions, deletions, and / or insertions of one or more amino acid residues. The method according to any one of claims 61-66 is characterized in that, The modified IL20RA protein has an amino acid sequence comprising the amino acid sequence shown in SEQ ID NO: 91; or, comprising an amino acid sequence with at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in SEQ ID NO: 91; or, comprising an amino acid sequence differing from the amino acid sequence shown in SEQ ID NO: 91 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprising an amino acid sequence similar to that shown in SEQ ID NO: 91, including substitutions, deletions, and / or insertions of one or more amino acid residues. The method according to any one of claims 61-67 is characterized in that, The nucleotide sequence encoding the human IL20RA region includes a portion of exon 2, the entirety of exon 3, the entirety of exon 4, the entirety of exon 5, and a portion of exon 6 of the human IL20RA gene. The method according to any one of claims 61-68, characterized in that, The nucleotide sequence encoding the human IL20RA region comprises the nucleotide sequence shown in positions 137004735-137017092 of NCBI Gene Accession Number NC_000006.12; or, comprises a nucleotide sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, or 98% identity to the nucleotide sequence shown in positions 137004735-137017092 of NCBI Gene Accession Number NC_000006.

12. Or at least 99% of the nucleotide sequence; or, a nucleotide sequence that differs from the nucleotide sequence shown in positions 137004735-137017092 of NCBI Gene Accession Number NC_000006.12 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2 or no more than 1 nucleotide; or, a nucleotide sequence that includes substitutions, deletions and / or insertions of one or more nucleotides as shown in the nucleotide sequence shown in positions 137004735-137017092 of NCBI Gene Accession Number NC_000006.

12. The method according to any one of claims 61-69 is characterized in that, The mRNA transcribed from the nucleotide sequence encoding the modified IL20RA protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 90; B) the nucleotide sequence being at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identical to the nucleotide sequence shown in SEQ ID NO: 90; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 90 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 90, including substitutions, deletions, and / or insertions of one or more nucleotides. The method according to any one of claims 61-70 is characterized in that, The expression of the modified IL20RA protein is controlled by endogenous IL20RA regulatory elements (e.g., promoter, 5'UTR and / or 3'UTR). The method according to any one of claims 61-71 is characterized in that, The non-human animal or non-human animal genome is homozygous or heterozygous for the genetically modified endogenous IL20RA gene. The method according to any one of claims 61-72 is characterized in that, The non-human animal or the genome of the non-human animal contains an endogenous gene encoding a human or modified other gene, preferably, the other gene is selected from at least one of IL22, IL22RA1, IL10RB, IL20RB, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3 and OX40. A non-human animal genome, characterized in that, The non-human animal or the non-human animal genome contains a genetically modified endogenous IL20RB gene encoding a modified IL20RB protein, and the non-human animal expresses the modified IL20RB protein. The non-human animal or non-human animal genome according to claim 74 is characterized in that, The non-human animal or non-human animal genome contains a nucleotide sequence that replaces the corresponding nucleotide sequence encoding the non-human animal endogenous IL20RB region with a nucleotide sequence encoding the human IL20RB region. The non-human animal or non-human animal genome according to any one of claims 74-75 is characterized in that, The modified IL20RB protein comprises a cytoplasmic domain, a transmembrane domain, and an extracellular domain; Preferably, the modified IL20RB protein contains an extracellular domain substantially the same as that of the human IL20RB protein; Preferably, all or part of the extracellular domain comprises the amino acid sequence shown in positions 44-233, 30-233, or 35-233 of SEQ ID NO: 113; or, comprises an amino acid sequence with an identity of at least 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with ...10, 9, 8, 7, 6, 5, 4, 3, 2, or at least 1 amino acid with the amino acid sequence shown in positions 44-233, 30-233, or 35-233 of SEQ ID NO: 113; or, comprises an amino acid sequence with an identity of at least 10, 9, 8, 7, 6, 5, 4, 3, 2, or at least 1 amino acid with the amino acid sequence shown in positions 44-233, 30-233, or 35-233 of SEQ ID NO: 113; or, comprises an amino acid sequence with an identity of at least 10, 9, 8, 7, 6, 5, 4, 3, 2, or at least 1 amino acid with the amino acid sequence shown The amino acid sequence shown in NO:113 at positions 44-233, 30-233, or 35-233, including substitutions, deletions, and / or insertions of one or more amino acid residues. The non-human animal or non-human animal genome according to any one of claims 74-76 is characterized in that, The modified IL20RB protein contains substantially the same transmembrane domain as the human IL20RB protein; Preferably, all or part of the transmembrane domain comprises the amino acid sequence shown at positions 234-250, 234-244, or 234-254 of SEQ ID NO: 113; or, comprises an amino acid sequence with an identity of at least 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with ...10, 9, 8, 7, 6, 5, 4, 3, 2, or at least 1 amino acid with the amino acid sequence shown at positions 234-250, 234-244, or 234-254 of SEQ ID NO: 113; or, comprises an amino acid sequence with an identity of at least 10, 9, 8, 7, 6, 5, 4, 3, 2, or at least 1 amino acid with the amino acid sequence shown at positions 234-250, 234-244, or 234-254 of SEQ ID NO: 113; or, comprises an amino acid sequence with an identity of at least 10, 9, 8, 7, 6, 5, 4, NO: 113 indicates the amino acid sequence represented by positions 234-250, 234-244, or 234-254, including substitutions, deletions, and / or insertions of one or more amino acid residues. The non-human animal or non-human animal genome according to any one of claims 74-77 is characterized in that, The nucleotide sequence encoding the human IL20RB region comprises the amino acid sequence shown in positions 44-244 of SEQ ID NO: 113; or comprises an amino acid sequence with an identity of at least 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown in positions 44-244 of SEQ ID NO: 113; or comprises an amino acid sequence differing from the amino acid sequence shown in positions 44-244 of SEQ ID NO: 113 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or comprises an amino acid sequence with an identity of positions 44-244 of SEQ ID NO: 113, including substitutions, deletions, and / or insertions of one or more amino acid residues. The non-human animal or non-human animal genome according to any one of claims 74-78 is characterized in that, The non-human animal mentioned is a rodent, preferably a rat or mouse; Preferably, the modified IL20RB protein contains a cytoplasmic domain that is substantially the same as that of the non-human endogenous IL20RB protein. Preferably, the amino acid sequence of the cytoplasmic domain comprises the amino acid sequence shown in positions 250-308 of SEQ ID NO: 112; or, comprises an amino acid sequence with at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in positions 250-308 of SEQ ID NO: 112; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 250-308 of SEQ ID NO: 112 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with positions 250-308 of SEQ ID NO: 112, including substitutions, deletions, and / or insertions of one or more amino acid residues. The non-human animal or non-human animal genome according to any one of claims 74-79 is characterized in that, The modified IL20RB protein has an amino acid sequence comprising the amino acid sequence shown in SEQ ID NO: 121; or, comprising an amino acid sequence with at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in SEQ ID NO: 121; or, comprising an amino acid sequence differing from the amino acid sequence shown in SEQ ID NO: 121 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprising an amino acid sequence similar to that shown in SEQ ID NO: 121, including substitutions, deletions, and / or insertions of one or more amino acid residues. The non-human animal or non-human animal genome according to any one of claims 74-80 is characterized in that, The nucleotide sequence encoding the human IL20RB region includes a portion of exon 2, all of exon 3, all of exon 4, all of exon 5, and a portion of exon 6 of the human IL20RB gene. The non-human animal or non-human animal genome according to any one of claims 74-81 is characterized in that, The nucleotide sequence encoding the human IL20RB region comprises the nucleotide sequence shown in positions 136980507-136995463 of NCBI Gene Accession Number NC_000003.12; or, comprises a nucleotide sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, or 98% identity to the nucleotide sequence shown in positions 136980507-136995463 of NCBI Gene Accession Number NC_000003.

12. Or at least 99% of the nucleotide sequence; or, a nucleotide sequence that differs from the nucleotide sequence shown in positions 136980507-136995463 of NCBI Gene Accession Number NC_000003.12 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2 or no more than 1 nucleotide; or, a nucleotide sequence that includes substitutions, deletions and / or insertions of one or more nucleotides as shown in the nucleotide sequence shown in positions 136980507-136995463 of NCBI Gene Accession Number NC_000003.

12. The non-human animal or non-human animal genome according to any one of claims 74-82 is characterized in that, The mRNA transcribed from the nucleotide sequence encoding the modified IL20RB protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 120; B) the nucleotide sequence being at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identical to the nucleotide sequence shown in SEQ ID NO: 120; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 120 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 120, including substitutions, deletions, and / or insertions of one or more nucleotides. The non-human animal or non-human animal genome according to any one of claims 74-83 is characterized in that, The expression of the modified IL20RB protein is controlled by endogenous IL20RB regulatory elements (e.g., promoter, 5'UTR and / or 3'UTR). The non-human animal or non-human animal genome according to any one of claims 74-84 is characterized in that, The non-human animal or non-human animal genome is homozygous or heterozygous for the genetically modified endogenous IL20RB gene. The non-human animal or non-human animal genome according to any one of claims 74-85 is characterized in that, The non-human animal or non-human animal genome contains endogenous genes encoding human or modified other genes, preferably, the other genes are selected from at least one of IL22, IL22RA1, IL10RB, IL20RA, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3 and OX40. A method for preparing non-human animals, characterized in that, The non-human animal contains a genetically modified endogenous IL20RB gene encoding a modified IL20RB protein, and the non-human animal expresses the modified IL20RB protein. The method according to claim 87, characterized in that, The genome of the non-human animal or non-human animal contains a nucleotide sequence that replaces the nucleotide sequence encoding the corresponding segment of the non-human animal endogenous IL20RB with a nucleotide sequence encoding the human IL20RB segment. The method according to any one of claims 87-88 is characterized in that, The modified IL20RB protein comprises a cytoplasmic domain, a transmembrane domain, and an extracellular domain; Preferably, the modified IL20RB protein contains an extracellular domain substantially the same as that of the human IL20RB protein; Preferably, all or part of the extracellular domain comprises the amino acid sequence shown in positions 44-233, 30-233, or 35-233 of SEQ ID NO: 113; or, comprises an amino acid sequence with an identity of at least 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with ...10, 9, 8, 7, 6, 5, 4, 3, 2, or at least 1 amino acid with the amino acid sequence shown in positions 44-233, 30-233, or 35-233 of SEQ ID NO: 113; or, comprises an amino acid sequence with an identity of at least 10, 9, 8, 7, 6, 5, 4, 3, 2, or at least 1 amino acid with the amino acid sequence shown in positions 44-233, 30-233, or 35-233 of SEQ ID NO: 113; or, comprises an amino acid sequence with an identity of at least 10, 9, 8, 7, 6, 5, 4, 3, 2, or at least 1 amino acid with the amino acid sequence shown The amino acid sequence shown in NO:113 at positions 44-233, 30-233, or 35-233, including substitutions, deletions, and / or insertions of one or more amino acid residues. The method according to any one of claims 87-89, characterized in that, The modified IL20RB protein contains substantially the same transmembrane domain as the human IL20RB protein; Preferably, all or part of the transmembrane domain comprises the amino acid sequence shown at positions 234-250, 234-244, or 234-254 of SEQ ID NO: 113; or, comprises an amino acid sequence with an identity of at least 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with ...10, 9, 8, 7, 6, 5, 4, 3, 2, or at least 1 amino acid with the amino acid sequence shown at positions 234-250, 234-244, or 234-254 of SEQ ID NO: 113; or, comprises an amino acid sequence with an identity of at least 10, 9, 8, 7, 6, 5, 4, 3, 2, or at least 1 amino acid with the amino acid sequence shown at positions 234-250, 234-244, or 234-254 of SEQ ID NO: 113; or, comprises an amino acid sequence with an identity of at least 10, 9, 8, 7, 6, 5, 4, NO: 113 indicates the amino acid sequence represented by positions 234-250, 234-244, or 234-254, including substitutions, deletions, and / or insertions of one or more amino acid residues. The method according to any one of claims 87-90, characterized in that, The nucleotide sequence encoding the human IL20RB region comprises the amino acid sequence shown in positions 44-244 of SEQ ID NO: 113; or comprises an amino acid sequence with an identity of at least 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown in positions 44-244 of SEQ ID NO: 113; or comprises an amino acid sequence differing from the amino acid sequence shown in positions 44-244 of SEQ ID NO: 113 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or comprises an amino acid sequence with an identity of positions 44-244 of SEQ ID NO: 113, including substitutions, deletions, and / or insertions of one or more amino acid residues. The method according to any one of claims 87-91, characterized in that, The non-human animal mentioned is a rodent, preferably a rat or mouse; Preferably, the modified IL20RB protein contains a cytoplasmic domain that is substantially the same as that of the non-human endogenous IL20RB protein. Preferably, the amino acid sequence of the cytoplasmic domain comprises the amino acid sequence shown in positions 250-308 of SEQ ID NO: 112; or, comprises an amino acid sequence with at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in positions 250-308 of SEQ ID NO: 112; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 250-308 of SEQ ID NO: 112 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with positions 250-308 of SEQ ID NO: 112, including substitutions, deletions, and / or insertions of one or more amino acid residues. The method according to any one of claims 87-92 is characterized in that, The modified IL20RB protein has an amino acid sequence comprising the amino acid sequence shown in SEQ ID NO: 121; or, comprising an amino acid sequence with at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in SEQ ID NO: 121; or, comprising an amino acid sequence differing from the amino acid sequence shown in SEQ ID NO: 121 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprising an amino acid sequence similar to that shown in SEQ ID NO: 121, including substitutions, deletions, and / or insertions of one or more amino acid residues. The method according to any one of claims 87-93 is characterized in that, The nucleotide sequence encoding the human IL20RB region includes a portion of exon 2, all of exon 3, all of exon 4, all of exon 5, and a portion of exon 6 of the human IL20RB gene. The method according to any one of claims 87-94, characterized in that, The nucleotide sequence encoding the human IL20RB region comprises the nucleotide sequence shown in positions 136980507-136995463 of NCBI Gene Accession Number NC_000003.12; or, comprises a nucleotide sequence with at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, or 98% identity to the nucleotide sequence shown in positions 136980507-136995463 of NCBI Gene Accession Number NC_000003.

12. Or at least 99% of the nucleotide sequence; or, a nucleotide sequence that differs from the nucleotide sequence shown in positions 136980507-136995463 of NCBI Gene Accession Number NC_000003.12 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2 or no more than 1 nucleotide; or, a nucleotide sequence that includes substitutions, deletions and / or insertions of one or more nucleotides as shown in the nucleotide sequence shown in positions 136980507-136995463 of NCBI Gene Accession Number NC_000003.

12. The method according to any one of claims 87-95, characterized in that, The mRNA transcribed from the nucleotide sequence encoding the modified IL20RB protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 120; B) the nucleotide sequence being at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identical to the nucleotide sequence shown in SEQ ID NO: 120; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 120 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 120, including substitutions, deletions, and / or insertions of one or more nucleotides. The method according to any one of claims 87-96, characterized in that, The expression of the modified IL20RB protein is controlled by endogenous IL20RB regulatory elements (e.g., promoter, 5'UTR and / or 3'UTR). The method according to any one of claims 87-97, characterized in that, The non-human animal or non-human animal genome is homozygous or heterozygous for the genetically modified endogenous IL20RB gene. The method according to any one of claims 87-98 is characterized in that, The non-human animal or the genome of the non-human animal contains an endogenous gene encoding a human or modified other gene. Preferably, the other gene is selected from at least one of IL22, IL22RA1, IL10RB, IL20RA, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3, and OX40. A non-human animal genome, characterized in that, The non-human animal or the non-human animal genome contains a genetically modified endogenous IL10RB gene encoding a modified IL10RB protein, and the non-human animal expresses the modified IL10RB protein. The non-human animal or non-human animal genome according to claim 100 is characterized in that, The non-human animal or non-human animal genome contains a nucleotide sequence that replaces the corresponding nucleotide sequence encoding the non-human animal endogenous IL10RB region with a nucleotide sequence encoding the human IL10RB region. The non-human animal or non-human animal genome according to any one of claims 100-101 is characterized in that, The modified IL10RB protein includes a cytoplasmic domain, a transmembrane domain, and an extracellular domain. Preferably, the modified IL10RB protein contains an extracellular domain substantially the same as that of the human IL10RB protein; Preferably, all or part of the extracellular domain comprises the amino acid sequence shown at positions 20-220 or 21-218 of SEQ ID NO: 57; or, comprises an amino acid sequence with an identity of at least 77%, 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown at positions 20-220 or 21-218 of SEQ ID NO: 57; or, comprises an amino acid sequence differing from the amino acid sequence shown at positions 20-220 or 21-218 of SEQ ID NO: 57 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with an amino acid sequence shown at positions 20-220 or 21-218 of SEQ ID NO: 57, including substitutions, deletions, and / or insertions of one or more amino acid residues. The non-human animal or non-human animal genome according to any one of claims 100-102 is characterized in that, The non-human animal is a rodent, more preferably a rat or mouse. The non-human animal or non-human animal genome according to any one of claims 100-103 is characterized in that, The modified IL10RB protein contains essentially the same cytoplasmic domain as the endogenous IL10RB protein in non-human animals. Preferably, the amino acid sequence of the cytoplasmic domain comprises the amino acid sequence shown in positions 244-351 of SEQ ID NO: 56; or, comprises an amino acid sequence with an identity of at least 77%, 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown in positions 244-351 of SEQ ID NO: 56; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 244-351 of SEQ ID NO: 56 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with an amino acid sequence shown in positions 244-351 of SEQ ID NO: 56, including substitutions, deletions, and / or insertions of one or more amino acid residues. The non-human animal or non-human animal genome according to any one of claims 100-104 is characterized in that, The modified IL10RB protein contains a transmembrane domain that is substantially the same as that of the non-human endogenous IL10RB protein. Preferably, the amino acid sequence of the transmembrane domain comprises the amino acid sequence shown in positions 223-243 of SEQ ID NO: 56; or, comprises an amino acid sequence with an identity of at least 77%, 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown in positions 223-243 of SEQ ID NO: 56; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 223-243 of SEQ ID NO: 56 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with an amino acid sequence shown in positions 223-243 of SEQ ID NO: 56, including substitutions, deletions, and / or insertions of one or more amino acid residues. The non-human animal or non-human animal genome according to any one of claims 100-105 is characterized in that, The modified IL10RB protein has an amino acid sequence comprising the amino acid sequence shown in SEQ ID NO: 62; or, comprising an amino acid sequence with at least 77%, 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in SEQ ID NO: 62; or, comprising an amino acid sequence differing from the amino acid sequence shown in SEQ ID NO: 62 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprising an amino acid sequence with the amino acid sequence shown in SEQ ID NO: 62, including substitutions, deletions, and / or insertions of one or more amino acid residues. The non-human animal or non-human animal genome according to any one of claims 100-106 is characterized in that, The nucleotide sequence encoding the human IL10RB region includes a portion of exon 2, the entirety of exon 3, the entirety of exon 4, the entirety of exon 5, and a portion of exon 6 of the human IL10RB gene. The non-human animal or non-human animal genome according to any one of claims 100-107 is characterized in that, The nucleotide sequence encoding the human IL10RB region comprises the nucleotide sequence or a variant thereof shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9, or comprises a nucleotide sequence or a variant thereof with an identity of at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, or 90% with the nucleotide sequence or a variant thereof shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.

9. 7%, 98%, or at least 99%; or, containing a nucleotide sequence that differs from the nucleotide sequence shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or a variant thereof by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or, containing a nucleotide sequence that includes substitutions, deletions, and / or insertions of one or more nucleotides as shown in the nucleotide sequence shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or a variant thereof. The variants include one or more of the following: C>T mutation at position 33276503, G>A mutation at position 33276818, or TTTGTGG>TTG mutation at positions 33277048-33277053. The non-human animal or non-human animal genome according to any one of claims 100-108 is characterized in that, The mRNA transcribed from the nucleotide sequence encoding the modified IL10RB protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 61; B) the nucleotide sequence being at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identical to the nucleotide sequence shown in SEQ ID NO: 61; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 61 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 61, including substitutions, deletions, and / or insertions of one or more nucleotides. The non-human animal or non-human animal genome according to any one of claims 100-109 is characterized in that, The expression of the modified IL10RB protein is controlled by endogenous IL10RB regulatory elements (e.g., promoter, 5'UTR and / or 3'UTR). The non-human animal or non-human animal genome according to any one of claims 100-110 is characterized in that, The non-human animal genome is homozygous or heterozygous for the genetically modified endogenous IL10RB gene. The non-human animal or non-human animal genome according to any one of claims 100-111 is characterized in that, The non-human animal or non-human animal genome contains endogenous genes encoding human or modified other genes, preferably, the other genes are selected from at least one of IL22, IL22RA, IL20RA, IL20RB, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3 and OX40. A method for preparing non-human animals, characterized in that, The non-human animal contains a genetically modified endogenous IL10RB gene encoding a modified IL10RB protein, and the non-human animal expresses the modified IL10RB protein. The method according to claim 113 is characterized in that, The non-human animal or the genome of the non-human animal contains a nucleotide sequence that replaces the nucleotide sequence encoding the corresponding segment of the non-human animal's endogenous IL10RB with a nucleotide sequence encoding the human IL10RB segment. The method according to any one of claims 113-114 is characterized in that, The modified IL10RB protein includes a cytoplasmic domain, a transmembrane domain, and an extracellular domain. Preferably, the modified IL10RB protein contains an extracellular domain substantially the same as that of the human IL10RB protein; Preferably, all or part of the extracellular domain comprises the amino acid sequence shown at positions 20-220 or 21-218 of SEQ ID NO: 57; or, comprises an amino acid sequence with an identity of at least 77%, 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown at positions 20-220 or 21-218 of SEQ ID NO: 57; or, comprises an amino acid sequence differing from the amino acid sequence shown at positions 20-220 or 21-218 of SEQ ID NO: 57 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with an amino acid sequence shown at positions 20-220 or 21-218 of SEQ ID NO: 57, including substitutions, deletions, and / or insertions of one or more amino acid residues. The method according to any one of claims 113-115 is characterized in that, The non-human animal is a rodent, more preferably a rat or mouse. The method according to any one of claims 113-116 is characterized in that, The modified IL10RB protein contains essentially the same cytoplasmic domain as the endogenous IL10RB protein in non-human animals. Preferably, the amino acid sequence of the cytoplasmic domain comprises the amino acid sequence shown in positions 244-351 of SEQ ID NO: 56; or, comprises an amino acid sequence with an identity of at least 77%, 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown in positions 244-351 of SEQ ID NO: 56; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 244-351 of SEQ ID NO: 56 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with an amino acid sequence shown in positions 244-351 of SEQ ID NO: 56, including substitutions, deletions, and / or insertions of one or more amino acid residues. The method according to any one of claims 113-117 is characterized in that, The modified IL10RB protein contains a transmembrane domain that is substantially the same as that of the non-human endogenous IL10RB protein. Preferably, the amino acid sequence of the transmembrane domain comprises the amino acid sequence shown in positions 223-243 of SEQ ID NO: 56; or, comprises an amino acid sequence with an identity of at least 77%, 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% with the amino acid sequence shown in positions 223-243 of SEQ ID NO: 56; or, comprises an amino acid sequence differing from the amino acid sequence shown in positions 223-243 of SEQ ID NO: 56 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprises an amino acid sequence with an amino acid sequence shown in positions 223-243 of SEQ ID NO: 56, including substitutions, deletions, and / or insertions of one or more amino acid residues. The method according to any one of claims 113-118 is characterized in that, The modified IL10RB protein has an amino acid sequence comprising the amino acid sequence shown in SEQ ID NO: 62; or, comprising an amino acid sequence with at least 77%, 78%, 79%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the amino acid sequence shown in SEQ ID NO: 62; or, comprising an amino acid sequence differing from the amino acid sequence shown in SEQ ID NO: 62 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 amino acid; or, comprising an amino acid sequence with the amino acid sequence shown in SEQ ID NO: 62, including substitutions, deletions, and / or insertions of one or more amino acid residues. The method according to any one of claims 113-119 is characterized in that, The nucleotide sequence encoding the human IL10RB region includes a portion of exon 2, the entirety of exon 3, the entirety of exon 4, the entirety of exon 5, and a portion of exon 6 of the human IL10RB gene. The method according to any one of claims 113-120 is characterized in that, The nucleotide sequence encoding the human IL10RB region comprises the nucleotide sequence or a variant thereof shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9, or comprises a nucleotide sequence or a variant thereof with an identity of at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, or 90% with the nucleotide sequence or a variant thereof shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.

9. 7%, 98%, or at least 99%; or, containing a nucleotide sequence that differs from the nucleotide sequence shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or a variant thereof by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or, containing a nucleotide sequence that includes substitutions, deletions, and / or insertions of one or more nucleotides as shown in the nucleotide sequence shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or a variant thereof. The variants include one or more of the following: C>T mutation at position 33276503, G>A mutation at position 33276818, or TTTGTGG>TTG mutation at positions 33277048-33277053. The method according to any one of claims 113-121 is characterized in that, The mRNA transcribed from the nucleotide sequence encoding the modified IL10RB protein comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 61; B) the nucleotide sequence being at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identical to the nucleotide sequence shown in SEQ ID NO: 61; C) the nucleotide sequence differing from the nucleotide sequence shown in SEQ ID NO: 61 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 61, including substitutions, deletions, and / or insertions of one or more nucleotides. The method according to any one of claims 113-122 is characterized in that, The expression of the modified IL10RB protein is controlled by endogenous IL10RB regulatory elements (e.g., promoter, 5'UTR and / or 3'UTR). The method according to any one of claims 113-123 is characterized in that, The non-human animal genome is homozygous or heterozygous for the genetically modified endogenous IL10RB gene. The method according to any one of claims 113-124 is characterized in that, The non-human animal or the genome of the non-human animal contains an endogenous gene encoding a human or modified or wild-type other gene, preferably, the other gene is selected from at least one of IL22, IL22RA, IL20RA, IL20RB, IL20, IL10, IL10RA, IL19, IL26, 4-1BB, PD-1, PD-L1, SIRPA, IL6, IL4, TIGIT, CTLA4, LAG3 and OX40. A nucleic acid, characterized in that, The nucleic acid comprises any one of the following groups: 1) A nucleotide sequence encoding the modified IL22RA1 protein as described in any one of claims 1-14; preferably, the nucleic acid further comprises SEQ ID NO: 30; preferably, the mRNA transcribed from the nucleic acid comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 31; B) the identity with the nucleotide sequence shown in SEQ ID NO: 31 is at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99%; C) the difference from the nucleotide sequence shown in SEQ ID NO: 31 is no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or, D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 31, including substitutions, deletions, and / or insertions of one or more nucleotides; 2) A nucleotide sequence encoding the human or modified IL22 protein as described in any one of claims 28-37; preferably, the nucleic acid further comprises SEQ ID NO: 5; preferably, the mRNA transcribed from the nucleic acid comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 6; B) the identity with the nucleotide sequence shown in SEQ ID NO: 6 is at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99%; C) the difference from the nucleotide sequence shown in SEQ ID NO: 6 is no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or, D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 6, including substitutions, deletions, and / or insertions of one or more nucleotides; 3) A nucleotide sequence encoding the modified IL20RA protein as described in any one of claims 48-60; preferably, the nucleic acid further comprises the nucleotide sequence shown at positions 137004735-137017092 of NCBI Gene Accession Number NC_000006.12; preferably, the mRNA transcribed from the nucleic acid comprises any one of the following: A) the nucleotide sequence shown in SEQ ID NO: 90; B) the identity with the nucleotide sequence shown in SEQ ID NO: 90 is at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99%; C) the difference from the nucleotide sequence shown in SEQ ID NO: 90 is no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or, D) has SEQ ID NO:

90. NO:90, a nucleotide sequence including substitutions, deletions, and / or insertions of one or more nucleotides; 4) A nucleotide sequence encoding the modified IL20RB protein as described in any one of claims 74-86; preferably, the nucleic acid further comprises the nucleotide sequence shown at positions 136980507-136995463 of NCBI Gene Accession Number NC_000003.12; preferably, the mRNA transcribed from the nucleic acid comprises any one of the following: A) the nucleotide sequence shown in SEQ ID NO: 120; B) at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% identity with the nucleotide sequence shown in SEQ ID NO: 120; C) differing from the nucleotide sequence shown in SEQ ID NO: 120 by no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) having SEQ ID NO:

120. The nucleotide sequence shown in NO: 120 includes nucleotide sequences with substitutions, deletions, and / or insertions of one or more nucleotides; 5) A nucleotide sequence encoding the modified IL10RB protein as described in any one of claims 100-112; preferably, the nucleic acid further comprises the nucleotide sequence shown at positions 33268405-33288111 of NCBI Gene Accession Number NC_000021.9 or a variant thereof; wherein the variant comprises one or more of the following: C>T mutation at position 33276503, G>A mutation at position 33276818, or TTTTGGG>TTG mutation at positions 33277048-33277053; Preferably, the mRNA transcribed from the nucleic acid comprises any one of the following groups: A) the nucleotide sequence shown in SEQ ID NO: 61; B) the identity with the nucleotide sequence shown in SEQ ID NO: 61 is at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99%; C) the difference from the nucleotide sequence shown in SEQ ID NO: 61 is no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, or no more than 1 nucleotide; or D) a nucleotide sequence having the nucleotide sequence shown in SEQ ID NO: 61, including substitutions, deletions, and / or insertions of one or more nucleotides. A polypeptide, characterized in that, The polypeptide includes any one of the following: 1) Encoded by the genetically modified endogenous IL22RA1 gene as described in any one of claims 1-14; 2) Encoded by the genetically modified endogenous IL22 gene as described in any one of claims 28-37; 3) Encoded by the genetically modified endogenous IL20RA gene as described in any one of claims 48-60; 4) Encoded by the genetically modified endogenous IL20RB gene as described in any one of claims 74-86; 5) Encoded by the genetically modified endogenous IL10RB gene as described in any one of claims 100-112; Preferably, the polypeptide is encoded by the nucleic acid of claim 126. A cell, tissue, or organ characterized by, The cells, tissues, or organs described herein are derived from any of the non-human animals described in claims 1-14, 28-37, 48-60, 74-86, and 100-112, or from non-human animals prepared by any of the methods described in claims 15-27, 38-47, 61-73, 87-99, and 113-125, and / or the cells, tissues, or organs contain the genome of any of the non-human animals described in claims 1-14, 28-37, 48-60, 74-86, and 100-112, and / or the cells, tissues, or organs contain the nucleic acid described in claim 126, and / or the cells, tissues, or organs express the polypeptide described in claim 127. The application of the polypeptide of claim 127, the nucleic acid of claim 126, the non-human animal or non-human animal genome of any one of claims 1-14, 28-37, 48-60, 74-86, 100-112, the non-human animal prepared by the method of any one of claims 15-27, 38-47, 61-73, 87-99, 113-125, and the cell, tissue or organ of claim 128, characterized in that, The applications include: A) Applications in product development involving immune processes in human cells related to IL22, IL22RA1, IL10RB, IL10, IL10RA, IL20RA and / or IL20RB; B) Application as a model system related to IL22, IL22RA1, IL10RB, IL10, IL10RA, IL20RA and / or IL20RB in pharmacological, immunological, microbiological and medical research; C) Involving the production and use of animal experimental disease models for etiological studies and / or for the development of diagnostic and / or therapeutic strategies related to IL22, IL22RA1, IL10RB, IL10, IL10RA, IL20RA and / or IL20RB; D) Its application in the screening, efficacy testing, evaluation, validation, or assessment of modulators of the human IL22, IL22RA1, IL10RB, IL10, IL10RA, IL20RA, and / or IL20RB signaling pathways in vivo; or, E) Investigate the gene functions of IL22, IL22RA1, IL10RB, IL20RA and / or IL20RB, investigate drugs and their efficacy targeting human IL22, IL22RA1, IL10RB, IL20RA and / or IL20RB, and investigate the application of drugs in the treatment of diseases related to IL22, IL22RA1, IL10RB, IL10, IL10RA, IL20RA and / or IL20RB.

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