Efficient bi-allelic gene knockout and site-specific knock-in mediated by TALENs in pigs

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	Efficient bi-allelic gene knockout and site-specific knock-in mediated by TALENs in pigs; Efficient bi-allelic gene knockout and site-specific knock-in mediated by TALENs in pigs
	Yao Jing ; HUANG Jiao-jiao ; HAI Tang ; WANG Xian-long ; QIN Guo-song ; ZHANG Hong-yong ; WU Rong ; CAO Chun-wei ; XI Jian-zhong ; ZHOU Qi ; YUAN Zeng-qiang ; ZHAO Jian-guo
	2016
关键词	TALENs GGTA1 Parkin DJ-1 Pig TALENs GGTA1 Parkin DJ-1 Pig
英文摘要	Objective To establish the platform of TALENs mediated gene editing in pigs and produce DJ-1 gene knockout(KO) pigs.Methods TALENs against GGTA1,Parkin and DJ-1 genes were co-transfected respectively into porcine fibroblast cells with pCAG-GFP plasmid to obtain gene KO colonies.And single-strand oligodeoxynucleotides(ssODN) ontaining Hindlll restriction enzyme site was co-transfected with TALENs against DJ-1 gene to test the possibility of TALEN-mediated site-specific knockin.Single colonies were screened by FACS,and targeted cell colonies were identified by PCR,enzyme digestion and sequencing.Further,mixed DJ-1 mutant colonies were used as donor cells for somatic cell nuclear transfer(SCNT) to produce DJ-1 KO pigs.Results In total,5%(2/40),2.5%(2/80),and 22%(11/50) of the obtained colonies of fibroblast cells were mutated for GGTA1,Parkin,and DJ-1,respectively.Among these mutant colonies,over 1/3were bi-allelic KO,and no off-target cleavage was detected.One of the ten mutant colonies had the Hindlll site ins...; Objective To establish the platform of TALENs mediated gene editing in pigs and produce DJ-1 gene knockout(KO) pigs.Methods TALENs against GGTA1,Parkin and DJ-1 genes were co-transfected respectively into porcine fibroblast cells with pCAG-GFP plasmid to obtain gene KO colonies.And single-strand oligodeoxynucleotides(ssODN) ontaining Hindlll restriction enzyme site was co-transfected with TALENs against DJ-1 gene to test the possibility of TALEN-mediated site-specific knockin.Single colonies were screened; 中国实验动物学会; 9
会议录	第十二届中国实验动物科学年会（2016.南宁）
语种	英语
内容类型	会议论文
源URL	[http://ir.pku.edu.cn/handle/20.500.11897/479852]
专题	工学院
推荐引用方式 GB/T 7714	Yao Jing,HUANG Jiao-jiao,HAI Tang,et al. Efficient bi-allelic gene knockout and site-specific knock-in mediated by TALENs in pigs, Efficient bi-allelic gene knockout and site-specific knock-in mediated by TALENs in pigs[C]. 见:.