Construction method and application of IL1RAP gene humanized non-human animal
A non-human animal, humanized technology, applied in the field of animal genetic engineering and genetic modification, biomedicine, can solve the problem of successful construction of humanized animals without IL1RAP gene, and no disclosure of exogenous nucleotide sequence IL1RAP gene human source issues such as transformation
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Embodiment 1I
[0236] Example 1IL1RAP gene humanized mice
[0237] Mouse IL1RAP gene (NCBI Gene ID: 16180, Primary source: MGI: 104975, UniProt: Q61730, located at positions 26400259 to 26548878 of chromosome 16 NC_000082.7, based on transcript NM_008364.2 and its encoded protein NP_032390.1 ( SEQ ID NO: 1)) and human IL1RAP gene (NCBI Gene ID: 3556, Primary source: HGNC: 5995, UniProt ID: Q9NPH3, located at positions 190514085 to 190659750 of chromosome 3 NC_000003.12, based on transcript NM_001167929.2 The comparison schematic diagram of its encoded protein NP_001161401.1 (SEQ ID NO: 2)) is as follows figure 1 shown.
[0238] In order to achieve the purpose of the present invention, a nucleotide sequence encoding human IL1RAP protein can be introduced into the mouse endogenous IL1RAP locus, so that the mouse expresses human or humanized IL1RAP protein, and there are various alternative ways. For example, using gene editing techniques, sequences encoding the human signal peptide and extra...
Embodiment 2
[0267] Example 2 Preparation of double humanized or multiple double humanized mice
[0268] Using this method or the prepared IL1RAP gene humanized mouse model can also be used to prepare double or multiple humanized mouse models. For example, in the aforementioned Example 1, embryonic stem cells used for blastocyst microinjection can be selected from mice with other genetic modifications such as IL33, PD-L1, IL10, IL1B, IL1R1, IL13, IL17, CCR5, CCR8, etc., or , or on the basis of humanized IL1RAP mice, the mouse ES embryonic stem cell isolation and gene recombination targeting technology can be used to obtain IL1RAP and other gene-modified double-gene or multi-gene-modified mouse models. The IL1RAP mouse homozygote or heterozygote obtained by this method can also be mated with other gene-modified homozygous or heterozygous mice, and the offspring are screened. According to the law of Mendelian inheritance, there is a certain probability to obtain humanized IL1RAP. With other...
Embodiment 3
[0269] Embodiment 3 drug efficacy verification
[0270] The IL1RAP humanized mice prepared by this method can be used to evaluate the efficacy of drugs targeting human IL1RAP. For example, IL1RAP humanized mouse homozygotes were subcutaneously inoculated with mouse colon cancer cells MC38. After the tumor volume grew to about 100 mm3, it was divided into a control group or a treatment group according to the tumor volume. The treatment group randomly selected drugs targeting human IL1RAP. The control group was injected with an equal volume of normal saline or PBS. Regularly measure the tumor volume and weigh the body weight of the mice, and the in vivo safety and efficacy of the drug can be effectively assessed by comparing the changes in the body weight of the mice and the tumor size.
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