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A method for constructing a mia3 conditional knockout mouse model

A mouse model and construction method technology, applied in the field of genetic engineering, can solve the problem of not finding knockout mice, etc., and achieve the effect of low cost, time saving and cost saving

Active Publication Date: 2022-07-29
广东药康生物科技有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0007] At present, there are no relevant reports on the construction of Mia3 gene conditional knockout mice using CRISPR / Cas9 technology

Method used

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  • A method for constructing a mia3 conditional knockout mouse model
  • A method for constructing a mia3 conditional knockout mouse model
  • A method for constructing a mia3 conditional knockout mouse model

Examples

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Effect test

Embodiment 1

[0040] Example 1. Construction of Mia3 gene conditional knockout mouse model

[0041] This example uses CRISPR Cas9 technology to genetically modify the mouse Mia3 gene. After the mice with stable genetic inheritance are mated with mice expressing Cre recombinase, the flox mice are knocked out, and knocking out this region will lead to the destruction of the protein function . Unless otherwise specified, the mice used in this example are C57BL / 6J strain mice.

[0042] 1. Determine the conditional knockout region and sequence

[0043] According to the structure of the Mia3 gene, the Exon2 to Exon7 part of the Mia3-202 transcript sequence was used as the knockout region, which contained a 3557bp coding sequence. The sequence of the targeting vector is shown below, where capital letters underlined correspond to Loxp sites. Loxp 1 is inserted upstream of the second exon (Exon 2), and Loxp 2 is inserted downstream of the seventh exon (Exon 7).

[0044] Among them, the insertion...

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Abstract

The present invention relates to a method for constructing a Mia3 gene conditional knockout mouse model by using CRISPR / Cas9 technology, comprising: 1. designing sgRNAs targeting mouse-derived Mia3 genes Exon2-Exon7, 2. using in vitro transcription technology to obtain the above-mentioned sgRNAs, 3. Co-injection or co-electroporation of the targeting vector, the above-mentioned sgRNA and Cas9 protein into mouse fertilized eggs and other steps. Compared with the traditional ES targeting mouse model, the Mia3 gene conditional knockout mouse model constructed by the present invention for the first time has the characteristics of high efficiency, fastness, simplicity and low cost, and can regulate genes in specific tissues or specific times through Cre. to express.

Description

technical field [0001] The invention belongs to the field of genetic engineering, and in particular relates to a method for constructing a Mia3 conditional gene knockout mouse model and its application. Background technique [0002] CRISPR / Cas9 technology realizes specific DNA recognition by designing specific sgRNA, and completes cutting at the target position, and then completes the repair of the break through the DNA repair mechanism of the cell itself, thereby realizing the "editing" of the target gene. With the development and in-depth research of CRISPR / Cas9 technology, it has been widely used to construct transgenic model animals (mainly gene-edited mice), thus providing basic research on animal-level treatments for preclinical use. [0003] The pathogenesis of human diseases and the screening of effective therapeutic drugs require extensive preclinical trials. Due to ethical restrictions on the direct use of human cells and tissues for preclinical research, animal m...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/89C12N15/87C12N9/22C12N15/113C12N15/12A01K67/027C12Q1/6888C12N15/11A61K49/00
CPCC12N15/89C12N15/87C12N9/22C12N15/1135C07K14/4703A01K67/0276C12Q1/6888A61K49/0008C12N2310/20A01K2217/075A01K2217/15A01K2217/206A01K2227/105A01K2267/0331A01K2267/0375C12Q2600/124C12Q2600/156
Inventor 梁馨元王韬蒋余亭黎晓雯郑桂纯
Owner 广东药康生物科技有限公司
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