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Application of the modified cytosine base-editing in the cultured cells of bama minipig
Pan, Jia sheng1; Lin, Zi sheng2; Wen, Jian cong2; Guo, Jian feng2; Wu, Xia hui2; Liu, Ying ying2; Lai, Wen jun2; Liang, Qi ying2; Xie, Yong shi2; Chen, Yi rou2; Chen, Yi hong2; Yan, Ai fen1,2; Feng, Juan2; Liu, Lian2; Gong, Dao yuan2; Zhu, Xiang xing1,2; Lu, Jia hong3; Tang, Dong sheng1,2
2021-09-01
Source PublicationBiotechnology Letters
ISSN0141-5492
Volume43Issue:9Pages:1699-1714
Abstract

Bama minipig is a unique miniature swine bred from China. Their favorable characteristics include delicious meat, strong adaptability, tolerance to rough feed, and high levels of stress tolerance. Unfavorable characteristics are their low lean meat percentage, high fat content, slow growth rate, and low feed conversion ratio. Genome-editing technology using CRISPR/Cas9 efficiently knocked out the myostatin gene (MSTN) that has a negative regulatory effect on muscle production, effectively promoting pig muscle growth and increasing lean meat percentage of the pigs. However, CRISPR/Cas9 genome editing technology is based on random mutations implemented by DNA double-strand breaks, which may trigger genomic off-target effects and chromosomal rearrangements. The application of CRISPR/Cas9 to improve economic traits in pigs has raised biosafety concerns. Base editor (BE) developed based on CRISPR/Cas9 such as cytosine base editor (CBE) effectively achieve targeted modification of a single base without relying on DNA double-strand breaks. Hence, the method has greater safety in the genetic improvement of pigs. The aim of the present study is to utilize a modified CBE to generate MSTN-knockout cells of Bama minipigs. Our results showed that the constructed “all-in-one”-modified CBE plasmid achieved directional conversion of a single C·G base pair to a T·A base pair of the MSTN target in Bama miniature pig fibroblast cells. We successfully constructed multiple single-cell colonies of Bama minipigs fibroblast cells carrying the MSTN premature termination and verified that there were no genomic off-target effects detected. This study provides a foundation for further application of somatic cell cloning to construct MSTN-edited Bama minipigs that carry only a single-base mutation and avoids biosafety risks to a large extent, thereby providing experience and a reference for the base editing of other genetic loci in Bama minipigs.

KeywordBama Minipig Cytosine Base Editor Genetic Modification Genome-editing Myostatin
DOI10.1007/s10529-021-03159-1
URLView the original
Indexed BySCIE
Language英語English
WOS Research AreaBiotechnology & Applied Microbiology
WOS SubjectBiotechnology & Applied Microbiology
WOS IDWOS:000668030100001
Scopus ID2-s2.0-85108973903
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Cited Times [WOS]:2   [WOS Record]     [Related Records in WOS]
Document TypeJournal article
CollectionUniversity of Macau
Corresponding AuthorZhu, Xiang xing; Lu, Jia hong; Tang, Dong sheng
Affiliation1.Guangdong Provincial Key Laboratory of Animal Molecular Design and Precise Breeding, School of Life Science and Engineering, Foshan University, Foshan, 528225, China
2.Guangdong Provincial Engineering and Technology Research Center for Gene Editing, School of Medicine, Foshan University, Foshan, 528225, China
3.State Key Laboratory of Quality Research in Chinese Medicine, Institute of Chinese Medical Sciences, University of Macau, 9999078, Macao
Corresponding Author AffilicationInstitute of Chinese Medical Sciences
Recommended Citation
GB/T 7714
Pan, Jia sheng,Lin, Zi sheng,Wen, Jian cong,et al. Application of the modified cytosine base-editing in the cultured cells of bama minipig[J]. Biotechnology Letters,2021,43(9):1699-1714.
APA Pan, Jia sheng,Lin, Zi sheng,Wen, Jian cong,Guo, Jian feng,Wu, Xia hui,Liu, Ying ying,Lai, Wen jun,Liang, Qi ying,Xie, Yong shi,Chen, Yi rou,Chen, Yi hong,Yan, Ai fen,Feng, Juan,Liu, Lian,Gong, Dao yuan,Zhu, Xiang xing,Lu, Jia hong,&Tang, Dong sheng.(2021).Application of the modified cytosine base-editing in the cultured cells of bama minipig.Biotechnology Letters,43(9),1699-1714.
MLA Pan, Jia sheng,et al."Application of the modified cytosine base-editing in the cultured cells of bama minipig".Biotechnology Letters 43.9(2021):1699-1714.
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