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Construction method and application of mir-124 gene knockout mouse animal model

A gene knockout mouse, mir-124 technology, applied in biochemical equipment and methods, genetic engineering, microorganisms, etc., can solve the problem of low expression level, achieve short modeling time, low price, simple and easy method Effect

Active Publication Date: 2019-01-22
CENT SOUTH UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0008] miR-124 is the most expressed miRNA in the mammalian nervous system, accounting for 5%-48% of the total miRNA in the mammalian cerebral cortex, but the expression level in other tissues is extremely low

Method used

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  • Construction method and application of mir-124 gene knockout mouse animal model
  • Construction method and application of mir-124 gene knockout mouse animal model
  • Construction method and application of mir-124 gene knockout mouse animal model

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Embodiment Construction

[0032] Construction method of mouse animal model:

[0033] (1) Construction of sgRNA targeting miR-124-1, miR-124-2, miR-124-3 genes, step-by-step sequencing, the required time is about 45-60 days; then linearize and purify the DNA and transcribe in vitro into sgRNA; purification of sgRNA to a purity suitable for transgene injection takes 15 days; the sgRNA sequence of the miR-124-1 is shown in SEQ ID NO.1; the sgRNA sequence of the miR-124-2 is shown in SEQ ID Shown in NO.2; the sgRNA sequence of the miR-124-3 is shown in SEQ ID NO.3;

[0034](2) C57 / BL6 female mice were treated with PMSG, injected with hCG 46 hours later, mated with male mice in a cage, and the fertilized eggs were taken for microinjection the next day, and the sgRNA described in step (1) was mixed with Cas9 nuclease mRNA After in vitro transcription, it takes 10 days to inject into fertilized eggs, and it takes 30 days to transfer the surviving fertilized eggs into pseudopregnant mice. 20 days after birth...

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Abstract

The invention discloses miR-124 gene knockout murine animal models. The murine animal models refer to mice with miR-124-1, miR-124-2 and miR-124-3 genes knocked out. With the adoption of a Crispr-cas9 gene knockout technology, microRNA (micro ribonucleic acid) miR-124 genes with the highest expression quantity in the brains of the mice are knocked out. The obtained mice show obvious nervous system disease states such as reduction of locomotor activity, deterioration of learning and memorizing abilities, increase of soluble amyloid beta protein and the like. Therefore, the simple, reliable and economical animal models can be provided for research of nervous system disease pathology and screening of drugs for treating nervous system diseases.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a miR-124 gene knockout mouse animal model and its construction method and application. Background technique [0002] On the one hand, modern people's life is fast-paced and under great pressure. On the other hand, the quality of life has generally improved, and the average life expectancy has increased. Therefore, the incidence of various mental diseases, especially senile degenerative diseases, is rising rapidly. At present, the diagnosis and staging of nervous system diseases are troublesome, and there is a lack of effective treatment methods. The reason is that the etiology of this type of disease is complex, the pathogenesis is unknown, and the current research lacks a suitable animal model. Therefore, it is an urgent task to establish effective disease animal models. The gene knockout mice constructed in this patent all showed obvious neurological disease states ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/113A01K67/027C12N15/85
CPCA01K67/0276A01K2217/075A01K2227/105A01K2267/03C12N15/113C12N15/85C12N2310/10C12N2800/107C12N2800/80
Inventor 朱曲波李大力童建斌殷永佳
Owner CENT SOUTH UNIV
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