Il21 gene knockout mouse model as well as construction method and application thereof

A gene-knockout mouse, a technique for constructing methods, applied to other methods of inserting foreign genetic materials, pharmaceutical formulations, using microinjection methods, etc.

Pending Publication Date: 2021-04-16
成都药康生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] At present, there is no successful case of using CRISPR / Cas9 technology to construct an IL-21 gene knockout mouse model

Method used

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  • Il21 gene knockout mouse model as well as construction method and application thereof
  • Il21 gene knockout mouse model as well as construction method and application thereof
  • Il21 gene knockout mouse model as well as construction method and application thereof

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Embodiment 1

[0043] A method for constructing an Il21 gene knockout mouse model, comprising the following steps:

[0044]S1, RISPR / Cas9 technology, according to the Il21 gene information, the exon3-exon5 of the Il21-201 (ENSMUST00000029273.7) transcript is used as a knockout region, which contains a 258bp coding sequence; according to this region, it is determined that the C57BL / 6J mouse IL10 gene is to be treated The knockout specific target sites gRNA1 and gRNA2, the gene sequences of gRNA1 and gRNA2 are respectively shown in SEQ ID NO.1 and SEQ ID NO.2 or shown in Table 1;

[0045] Table 1 gRNA sequence information

[0046] sgRNA name gRNA sequence(5'*3') PAM sgRNA1 TGCCTATGTATGTGTCGTCC TGG sgRNA2 TGGAGTGCCCACCTATCTGA TGG

[0047] S2. Microinject active gRNA1, gRNA2, and cas9 proteins into mouse fertilized eggs, take the fertilized eggs that survived the injection and transplant them into pseudopregnant female mice, and obtain F0 generation mice afte...

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Abstract

The invention discloses an Il21 gene knockout mouse model and a construction method and an application thereof. The construction method comprises the following steps that S1, based on the CRISPR/Cas9 technology, a knockout area of an Il21 gene is determined, specific target sites sgRNA1 and sgRNA2 are designed according to the determined knockout area, and gene sequences of the gRNA1 and the gRNA2 are shown in SEQ ID NO.1 and SEQ ID NO.2 respectively; S2, microinjecting active gRNA1, gRNA2 and cas9 proteins into fertilized eggs of the mouse, transplanting the fertilized eggs survived after injection into the body of the false pregnant female mouse, and obtaining an F0-generation mouse after the false pregnant female mouse is pregnant and farrowing; S3, mating the F0-generation mouse obtained in the step S2 with a wild-type mouse to obtain an F1-generation heterozygote; and S4, carrying out inbreeding on the F1-generation heterozygote obtained in the step S3 to obtain F2-generation mice, namely the IL21 gene knockout mouse model. Based on the CRISPR/Cas9 technology, the mouse animal model is established by knocking out an exon3-exon5 region in an ENSMUST000029273.7 transcript of an IL21 gene for the first time by designing reasonable gene sequences of specific target sites gRNA1 and gRNA2, and the mouse animal model can be applied to related research after tumor healing.

Description

technical field [0001] The invention relates to the technical field of gene knockout mouse model construction, in particular to an Il21 gene knockout mouse model and its construction method and application. Background technique [0002] IL-21 is interleukin-21 (interleukin-21, IL-21) is a secretory protein secreted by activated CD4+ T cells, its molecular structure is similar to IL-15, and it belongs to the IL-15 / IL-21 family , which binds to a complex receptor consisting of a private receptor (IL21R) and a common cytokine receptor chain (gamma(C)). IL-21 affects multiple cell types including CD8+ memory T cells, NK cells and CD4 memory T cell subsets. Its receptors are widely distributed on lymphoid hematopoietic cells. IL-21 enhances the antigen-specific response of immune cells, promotes the anti-tumor activity of CD8+ T cells and NK cells, and plays a role by binding to the specific type I cytokine receptor IL-21R. Previous studies have shown that IL-21 deficiency can...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/113C12N15/85C12N15/89C12N5/10A01K67/027A01K67/02A61K49/00
Inventor 史培良樊皓月辛闻婷
Owner 成都药康生物科技有限公司
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