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African green monkey kidney cell line capable of stably expressing SLAM protein as well as construction method and application of African green monkey kidney cell line

A green monkey kidney cell, stable expression technology, applied in artificial cell constructs, genetically modified cells, urinary tract/kidney cells, etc. simple effect

Pending Publication Date: 2022-03-22
INST OF ANIMAL HEALTH GUANGDONG ACADEMY OF AGRI SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] As an acute and severe infectious disease, CDV is extremely harmful because there is no specific drug treatment

Method used

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  • African green monkey kidney cell line capable of stably expressing SLAM protein as well as construction method and application of African green monkey kidney cell line
  • African green monkey kidney cell line capable of stably expressing SLAM protein as well as construction method and application of African green monkey kidney cell line
  • African green monkey kidney cell line capable of stably expressing SLAM protein as well as construction method and application of African green monkey kidney cell line

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Example 1 Construction of the African green monkey kidney cell line Vero-E6-SLAM stably expressing SLAM protein

[0042] (1) Test material

[0043] African green monkey kidney cells (Vero-E6) were purchased from Wuhan Punuosheng Life Science and Technology Co., Ltd., and were preserved in our laboratory; Genome-CRISPR TM Human AVVS1 Safe Harbor gene knock-in kit (pAAVS1-Doner, pCas-Guide-AAVS1 plasmid) was purchased from Guangzhou Yijin Biotechnology Co., Ltd. and kept by our laboratory; plasmid 1 (pUC57-SLAM) was purchased from Guangzhou Qingke Biotechnology Co., Ltd. Synthesized by Science and Technology Co., Ltd., its plasmid map is as follows figure 1 shown.

[0044] (2) Design of primers

[0045] According to the gene sequence of SLAM (NCBI accession number is EU678639.1), primers were designed, and the 5' ends of the upstream and downstream primers were respectively connected with the enzyme cutting sites XbaI and SwaI and the corresponding protection bases (se...

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Abstract

The invention belongs to the technical field of gene recombination and protein expression, and particularly relates to an African green monkey kidney cell line capable of stably expressing SLAM protein as well as a construction method and application of the African green monkey kidney cell line, the cell line is African green monkey kidney cells Vero-E6-SLAM, the African green monkey kidney cells Vero-E6-SLAM are sent to Guangdong Microbial Culture Collection Center on November 11, 2021 to be preserved, and the preservation number is GDMCCNO: 62054. The preparation method comprises the following steps: constructing a pAAVS1-Donor-SLAM overexpression plasmid, and then co-transfecting the pAAVS1-Donor-SLAM overexpression plasmid and pCass-Guide-AAVS1 into a Vero-E6 cell, so as to obtain the recombinant vector. According to the cell line, an exogenous gene can be integrated into a known safe site in a genome, normal expression of the transferred gene is realized without influencing the normal physiological function of the cell, the cloning accuracy is high, the cell line can be applied to high-yield SLAM protein, and a platform is provided for separation and pathogenic mechanism research of canine distemper virus.

Description

technical field [0001] The invention belongs to the technical field of gene recombination and protein expression, and in particular relates to an African green monkey kidney cell line stably expressing SLAM protein and its construction method and application. Background technique [0002] Canine distemper (Canine distemper, CD) is an acute, febrile and highly contagious infectious disease caused by canine distemper virus (Canine distempervirus, CDV) of the genus Morbillivirus in the Paramyxoviridae family. The main clinical features are biphasic fever, mucous membrane inflammation of the eyes, nose, and digestive tract, and catarrhal pneumonia, dermatitis, and neurological symptoms. Since it was first reported in 1809, the disease has become a worldwide epidemic, seriously affecting companion animals and wildlife. [0003] As an acute and severe infectious disease, CDV is extremely harmful because there is no specific drug treatment. The currently known CDV hosts have been...

Claims

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Application Information

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IPC IPC(8): C12N5/071C12N5/10C12N15/85C07K14/705
CPCC12N5/0686C12N15/85C07K14/70503C12N2510/02
Inventor 陈晶王晓虎张翩向华黄元黄忠王刚
Owner INST OF ANIMAL HEALTH GUANGDONG ACADEMY OF AGRI SCI
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