Rapid screening of single guide RNA targeting pig genome and the harvesting of monoclonal cells by microarray seal

The emergence of regular short repetitive palindromic sequence clusters (CRISPR) and CRISPR- associated proteins 9 (Cas9) gene editing technology has greatly promoted the wide application of genetically modified pigs. Efficient single guide RNA (sgRNA) is the key to the success of gene editing using...

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Bibliographic Details
Published in:Sheng wu yi xue gong cheng xue za zhi 2021-02, Vol.38 (1), p.111-121
Main Authors: Gao, Mengyu, Zhu, Xinglong, Wang, Shisheng, Zhang, Bingqi, Zhang, Yunlin, He, Yuting, Zhou, Yanyan, Li, Shun, Yang, Guang, Liao, Guangneng, Bao, Ji, Bu, Hong
Format: Article
Language:Chinese
Subjects:
Animals
Bioreactors
Cell culture
Clustered Regularly Interspaced Short Palindromic Repeats - genetics
CRISPR
CRISPR-Cas Systems - genetics
DNA microarrays
FAH gene
Gene Editing
Genetic modification
Genomes
Hogs
Reproductive cycle
Ribonucleic acid
RNA
RNA, Guide, CRISPR-Cas Systems - genetics
Screening
Swine
Technology utilization
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Summary:The emergence of regular short repetitive palindromic sequence clusters (CRISPR) and CRISPR- associated proteins 9 (Cas9) gene editing technology has greatly promoted the wide application of genetically modified pigs. Efficient single guide RNA (sgRNA) is the key to the success of gene editing using CRISPR/Cas9 technology. For large animals with a long reproductive cycle, such as pigs, it is necessary to screen out efficient sgRNA to avoid wasting time and resource costs before animal experiments. In addition, how to efficiently obtain positive gene editing monoclonal cells is a difficult problem to be solved. In this study, a rapid sgRNA screening method targeting the pig genome was established and we rapidly obtained gene edited cells, laying a foundation for the subsequent production of knockout pigs as human hepatocyte bioreactor. At the same time, the method of obtaining monoclonal cells using pattern microarray culture technology was explored.
ISSN:1001-5515
DOI:10.7507/1001-5515.202006032