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Construction method and application of Psm3 conditional gene knockout mouse model

A gene knockout mouse, the technology of construction method, applied in the field of medicine, to achieve the effect of reducing changes

Pending Publication Date: 2021-12-17
EAST CHINA NORMAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] However, the mechanism of Psme3 involved in tumorigenesis and development needs to be further elucidated

Method used

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  • Construction method and application of Psm3 conditional gene knockout mouse model
  • Construction method and application of Psm3 conditional gene knockout mouse model
  • Construction method and application of Psm3 conditional gene knockout mouse model

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Embodiment 1

[0079] 1. Construction of sgRNA. Using the Cas9 / RNA system gene targeting technology, a gRNA targeting the mouse Psme3 gene was constructed to guide the Cas9 protein to cut DNA double strands at the specific site of the crRNA guide sequence target; and the sgRNA cutting efficiency was tested in vitro.

[0080] 2. Design the corresponding donor according to the sgRNA, construct the donor plasmid, and make the Donorvector carrying the loxp fragment.

[0081] 3. Pronucleus injection and transplantation: superovulation of mice; injection of fertilized eggs, transplantation; obtaining F0 generation mice.

[0082] 4. Identification and breeding of F0 generation mice: positive mice were verified by PCR and sequencing; positive F0 generation mice were backcrossed with C57BL / 6J mice to obtain F1 generation mice.

[0083] 5. Identification of F1 generation mice: Positive mice were verified by PCR and sequencing.

[0084] 6. Genotype identification of F2 generation mice:

[0085] (1) ...

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Abstract

The invention discloses a construction method of a Psm3 conditional gene knockout mouse model and application of the Psm3 conditional gene knockout mouse model. According to the construction method, the third exon to the fifth exon of the Psm3 gene are knocked out by using a Cas9 / RNA system gene targeting technology, so that inactivation of the whole gene is realized, and the Psm3 gene conditional knockout mouse model is obtained. The Psme3 is used as a proteasome activating factor to regulate the hydrolytic activity of 20S proteasome and regulate and control the degradation of various tumor-related factors. The Psm3 gene conditional knockout mouse model constructed by the invention lays a foundation for further research on biological functions of the Psm3 gene, and can provide an ideal model for an action mechanism in occurrence and development of solid tumors with high expression of Psm3.

Description

technical field [0001] The invention belongs to the field of medicine, and in particular relates to a method for constructing a Psme3 conditional gene knockout mouse model and an application thereof. Background technique [0002] REGγ, also known as 11Sγ, PA28γ, PSME3 or Ki (GeneID: 10197), is a member of the 11S proteasome activator family. It was first discovered in the serum of patients with systemic lupus erythematosus and named as Ki antigen, but its The relationship with patients with systemic lupus erythematosus is still not very clear. Subsequent studies found that the Ki antigen, like REGα and REGβ, belongs to the 11S proteasome activator family member, and was named REGγ because it was discovered after REGα and REGβ. [0003] As a proteasome activating factor, REGγ binds to the α subunit of the 20S core particle, opens the 20S gate, and sends the substrate protein to the protease catalytic site of the β subunit of 20S, degrading the substrate protein. The REGγ-20...

Claims

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Application Information

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IPC IPC(8): C12N15/85C12N15/90C12N15/12A01K67/027C12N15/113C12N15/11
CPCC12N15/8509C12N15/907C07K14/4705A01K67/0276C12N15/113C12N2800/107C12N2800/30A01K2217/075A01K2227/105A01K2267/03C12N2310/20
Inventor 李晓涛沈诗慧李磊童璐高晓祝恩浩杜营营
Owner EAST CHINA NORMAL UNIV
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