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Construction method and application of non-human animal modified by humanized cell factor IL3 (interleukin3) gene

A non-human animal, genetic modification technology, applied in the direction of plant genetic improvement, chemical instruments and methods, botany equipment and methods, etc., can solve problems such as gene complexity

Active Publication Date: 2020-05-08
BIOCYTOGEN JIANGSU CO LTD +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, due to the differences in physiology and pathology between animals and humans, coupled with the complexity of genes (i.e., genetic factors), how to construct an "effective" humanized animal model for new drug development is still the biggest challenge ( 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)

Method used

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  • Construction method and application of non-human animal modified by humanized cell factor IL3 (interleukin3) gene
  • Construction method and application of non-human animal modified by humanized cell factor IL3 (interleukin3) gene
  • Construction method and application of non-human animal modified by humanized cell factor IL3 (interleukin3) gene

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1I

[0160] Example 1 IL3 Gene Humanized Mice

[0161] Mouse IL3 gene (NCBI Gene ID: 16187, Primary source: MGI: 96552, UniProt ID: P01586) (based on the transcript of NM_010556.4 → NP_034686.2, its mRNA sequence is shown in SEQ ID NO: 1, the corresponding The amino acid sequence is shown in SEQ ID NO: 2) and human IL3 gene (NCBI Gene ID: 3562, Primary source: HGNC: 6011, UniProt ID: P08700) (based on the transcript of NM_000588.3 → NP_000579.2, its mRNA sequence As shown in SEQ ID NO: 3, the corresponding amino acid sequence is shown in SEQ ID NO: 4) The comparison diagram is shown in figure 1 shown.

[0162] In order to achieve the purpose of the present invention, the gene sequence encoding human IL3 protein can be introduced into the endogenous mouse IL3 gene locus, so that the mouse expresses human IL3 protein. For example, gene editing technology can be used to modify mouse cells, insert the coding sequence that can express human IL3 protein after the endogenous mouse IL3 s...

Embodiment 2

[0211] Example 2 Generation of IL3 Humanized Cytokine Mice with Severe Immunodeficiency

[0212] In order to generate mice containing human IL3 and having severe immunodeficiency, the IL3 gene humanized heterozygous mice prepared in Example 1 can be mated with B-NDG mice or in vitro fertilized (IVF), according to Mendelian inheritance According to the law, the offspring can be screened with a certain probability to obtain heterozygous mice with IL3 humanization and IL-2Rγ chain deletion, and then cross the heterozygotes to obtain homozygotes with double genes or multiple genes.

[0213] In Example 1, the fertilized egg cells of B-NDG mice were used to replace NOD / scid mice in the step of microinjection, and B-NDG mice expressing human IL3 protein were directly obtained.

Embodiment 3

[0214] Example 3 Double-gene or multi-gene humanized mice containing human IL3

[0215] Double-humanized or multi-humanized mouse models can also be prepared by using the method or the prepared IL3 mice. For example, in the aforementioned Example 1, the fertilized egg cells used in the microinjection and embryo transfer process were selected from fertilized egg cells derived from other genetically modified mice, for example, the fertilized egg cells of CSF2 or IL15 or CSF1 gene humanized mice were selected according to this method. Methods Through gene editing, a double-gene humanized mouse model of CSF2 or IL15 or CSF1 gene humanization and IL3 gene modification can be further obtained.

[0216] The IL3 gene humanized mouse homozygous or heterozygous obtained by this method is mated or fertilized in vitro with other genetically modified homozygous or heterozygous mice, and the offspring are screened. According to the law of Mendelian inheritance, there may be a certain probab...

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Abstract

The invention relates to the modification of a non-human animal by a humanized gene, in particular to a construction method of a non-human animal modified by a humanized cell factor IL3 (interleukin3)gene, and application of the construction method in the biological medicine field, and in particular to an animal model which expresses a humanized IL3 protein. In certain examples, the genetic modification non-human animal which expresses the human IL3 protein also contains IL2rg deletion and / or contains the humanization of more cell factor, including CSF2 (colony stimulating factor 2) and thelike. The invention also provides a construction method for the above animal model and the application of the construction method in the biological medicine field.

Description

technical field [0001] This application relates to the establishment method and application of a humanized genetically modified animal model, specifically, to the construction method of a humanized cytokine IL3 protein-modified animal model and its application in biomedicine. Background technique [0002] The differentiation, development, proliferation and even activation of cells are all subject to the synergistic effect of multiple cytokine signals, among which interleukin 3 (interleukin3, IL3, IL-3) and granulocyte-macrophage colony stimulating factor (granulocyte-macrophage colony stimulating factor , GM-CSF, also known as colony stimulating factor 2, CSF2) These two cytokines play an important role in the development, differentiation and proliferation of myeloid cells. IL3 is a hematopoietic cell colony-stimulating factor (Granulocyte / macrophage colony-stimulating factors), which transmits growth and differentiation signals by binding to receptors on the surface of targ...

Claims

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Application Information

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IPC IPC(8): C12N15/24C12N15/85C12N5/10C12N15/113C07K19/00C12N15/62A01K67/027
CPCA01K67/0278A01K2217/072A01K2227/105A01K2267/0331A01K2267/0387C07K14/5403C07K2319/00C12N5/0602C12N15/113C12N2310/10C12N2510/00
Inventor 沈月雷郭朝设郭雅南白阳黄蕤尚诚彰张美玲姚佳维
Owner BIOCYTOGEN JIANGSU CO LTD
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