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CHO cell line capable of stably expressing African swine fever CD2v protein as well as construction method and application of CHO cell line

An African swine fever and stable expression technology, applied in the field of CHO cell line and its construction, can solve problems such as hindering the progress of research and lack of knowledge of ASF virus, and achieve high cloning accuracy, good repeatability, and simple construction process. Effect

Pending Publication Date: 2022-03-01
INST OF ANIMAL HEALTH GUANGDONG ACADEMY OF AGRI SCI
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AI Technical Summary

Problems solved by technology

A major reason for the difficulty of controlling the outbreak is that there is currently no commercially available effective vaccine worldwide, and the lack of knowledge about ASF virus (ASFV) antigens hinders the progress of related research

Method used

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  • CHO cell line capable of stably expressing African swine fever CD2v protein as well as construction method and application of CHO cell line
  • CHO cell line capable of stably expressing African swine fever CD2v protein as well as construction method and application of CHO cell line
  • CHO cell line capable of stably expressing African swine fever CD2v protein as well as construction method and application of CHO cell line

Examples

Experimental program
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Effect test

Embodiment 1

[0048] Example 1 Construction of CHO cell line CHO-CD2v stably expressing African swine fever CD2v protein

[0049] 1. Test material

[0050] Escherichia coli competent cells DH5α were purchased from Guangzhou Qingke Biotechnology Co., Ltd.; CHO cells were purchased from Wuhan Punuosai Life Technology Co., Ltd., and were preserved in our laboratory; mouse ROSA26 Safe Harbor gene knock-in vectors (including DC-SH02, pROSA26-Cas9 gRNA) was purchased from Guangzhou Yijin Biotechnology Co., Ltd. and preserved by our laboratory; fetal bovine serum, DMEM / F-12K medium, liposome LipofectamineTM 3000, puromycin, plasmid extraction kit, genome Kits such as DNA extraction and RNA extraction and Phusion Hot Start II High-Fidelity PCR MasterMix were purchased from Thermo Fisher Scientific (China) Co., Ltd.); restriction enzymes KpnI and BamHI and T4 DNA ligase were Purchased from NEB; the pMD18-CD2v plasmid carrying the CD2v gene was donated by Professor Hu Rongliang of the Academy of Mil...

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Abstract

The invention belongs to the technical field of cell recombination and protein expression, particularly relates to a CHO cell line for stably expressing African swine fever CD2v protein as well as a construction method and application of the CHO cell line, and discloses the CHO cell line for stably expressing the African swine fever CD2v protein, which is a Chinese hamster ovary cell CHO-CD2v. The strain is sent to Guangdong Microbial Culture Collection Center for preservation on November 11, 2021, and the preservation number is GDMCC No: 62052. According to the cell line, an exogenous gene CD2v is integrated into a known safe site in a Chinese hamster genome, so that normal expression of a transferred gene is realized without influencing normal physiological functions of cells, and the cell line is simple in construction process and high in cloning accuracy. The cell line can be used in the aspects of high-yield CD2v protein, drug effect screening of African swine fever, vaccine production and the like, and has very important significance in diagnosis of African swine fever and vaccine production.

Description

technical field [0001] The invention belongs to the technical field of cell recombination and protein expression, and in particular relates to a CHO cell line stably expressing African swine fever CD2v protein and its construction method and application. Background technique [0002] African swine fever (ASF) is a hemorrhagic, highly contagious disease of pigs caused by ASF virus (ASFV), and its clinical symptoms are very similar to those of swine fever. ASF is an emerging disease in the swine industry worldwide. The disease first occurred in Shenyang City, Liaoning Province, my country in August 2018. The introduction of ASF has seriously threatened the survival of my country's pig industry. One of the main reasons why the epidemic is difficult to control is that there is currently no commercially available effective vaccine worldwide, and the progress of related research is hindered by the lack of knowledge about ASF virus (ASFV) antigens. Studies have shown that ASFV se...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/071C12N5/10C12N15/34C12N15/86C12Q1/02A61K39/12A61P31/20C12R1/91
CPCC12N5/0682C07K14/005C12N15/86G01N33/5008A61K39/12A61P31/20C12N2510/02C12N2710/12022C12N2710/12034C12N2710/12043C12N2503/02A61K2039/5156A61K2039/53
Inventor 王晓虎陈晶王艳云向华黄元黄忠王刚
Owner INST OF ANIMAL HEALTH GUANGDONG ACADEMY OF AGRI SCI
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