Efficient Generation of Gene-Modified Pigs Harboring Precise Orthologous Human Mutation via CRISPR/Cas9-Induced Homology-Directed Repair in Zygotes

doi:10.1002/humu.22913

	Efficient Generation of Gene-Modified Pigs Harboring Precise Orthologous Human Mutation via CRISPR/Cas9-Induced Homology-Directed Repair in Zygotes
	Zhou, Xiaoyang 1; Wang, Lulu 1; Du, Yinan 2; Xie, Fei 1; Li, Liang 1; Liu, Yu 1; Liu, Chuanhong 1; Wang, Shiqiang 1; Zhang, Shibing 1; Huang, Xingxu2 ; Wang, Yong 1; Wei, Hong 1
	2016-01
发表期刊	HUMAN MUTATION
ISSN	1059-7794
卷号	37 期号:1 页码:110-118
发表状态	已发表
DOI	10.1002/humu.22913
摘要	Precise genetic mutation of model animals is highly valuable for functional investigation of human mutations. Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated 9 (Cas9)-induced homology-directed repair (HDR) is usually used for precise genetic mutation, being limited by the relatively low efficiency compared with that of non-homologous end joining (NHEJ). Although inhibition of NHEJ was shown to enhance HDR-derived mutation, in this work, without inhibition of NHEJ, we first generated gene-modified pigs harboring precise orthologous human mutation (Sox10 c.A325>T) via CRISPR/Cas9-induced HDR in zygotes using single-strand oligo DNA (ssODN) as template with an efficiency as high as 80%, indicating that pig zygotes exhibited high activities of HDR relative to NHEJ and were highly amendable to genetic mutation via CIRSPR/Cas9-induced HDR. Besides, we found a higher concentration of ssODN remarkably reduced HDR-derived mutation in pig zygotes, suggesting a possible balance for optimal HDR-derived mutation in zygotes between the excessive accessibility to HDR templates and the activities of HDR relative to NHEJ which appeared to be negatively correlated to ssODN concentration. In addition, the HDR-derived mutation, as well as those from NHEJ, extensively integrated into various tissues including gonad of founder pig without detected off-targeting, suggesting CRISPR/Cas9-induced HDR in zygotes is a reliable approach for precise genetic mutation in pigs.
关键词	SOX10 CRISPR Cas9 homology-directed repair pigs model organism disease model
收录类别	SCI
语种	英语
资助项目	National Major Scientific Projects Specifically for R&D of Transgenic Organisms[2014ZX08009-003-006]
WOS研究方向	Genetics & Heredity
WOS类目	Genetics & Heredity
WOS记录号	WOS:000369159900011
出版者	WILEY
WOS关键词	CONDITIONAL ALLELES ; CLONING EFFICIENCY ; KNOCKOUT RATS ; NEURAL CREST ; SOX10 ; SPECIFICITY ; CLEAVAGE
原始文献类型	Article
引用统计
文献类型	期刊论文
条目标识符	https://kms.shanghaitech.edu.cn/handle/2MSLDSTB/2053
专题	生命科学与技术学院_PI研究组_黄行许组
通讯作者	Wang, Yong; Wei, Hong
作者单位	1.Third Mil Med Univ, Coll Basic Med Sci, Dept Lab Anim Sci, Chongqing, Peoples R China 2.ShanghaiTech Univ, Sch Life Sci & Technol, Shanghai, Peoples R China
推荐引用方式 GB/T 7714	Zhou, Xiaoyang,Wang, Lulu,Du, Yinan,et al. Efficient Generation of Gene-Modified Pigs Harboring Precise Orthologous Human Mutation via CRISPR/Cas9-Induced Homology-Directed Repair in Zygotes[J]. HUMAN MUTATION,2016,37(1):110-118.
APA	Zhou, Xiaoyang.,Wang, Lulu.,Du, Yinan.,Xie, Fei.,Li, Liang.,...&Wei, Hong.(2016).Efficient Generation of Gene-Modified Pigs Harboring Precise Orthologous Human Mutation via CRISPR/Cas9-Induced Homology-Directed Repair in Zygotes.HUMAN MUTATION,37(1),110-118.
MLA	Zhou, Xiaoyang,et al."Efficient Generation of Gene-Modified Pigs Harboring Precise Orthologous Human Mutation via CRISPR/Cas9-Induced Homology-Directed Repair in Zygotes".HUMAN MUTATION 37.1(2016):110-118.