Targeted mutagenesis in the olive flounder (Paralichthys olivaceus) using the CRISPR/Cas9 system with electroporation

doi:10.2478/s11756-020-00677-7

CORC > 海洋研究所 > 中国科学院海洋研究所 > 实验海洋生物学重点实验室

	Targeted mutagenesis in the olive flounder (Paralichthys olivaceus) using the CRISPR/Cas9 system with electroporation
	Wang, Ling 1,2,4; Tan, Xungang 1,2; Wu, Zhihao 1,2; Wang, Lijuan 1,2; Jiao, Shuang 1,2; Zou, Yuxia 1,2; Ji, Guanglei 3,5; You, Feng 1,2
刊名	BIOLOGIA
	2021-01-20
页码	8
关键词	Genome editing CRISPR/Cas9 system Electroporation Myomaker and gsdf Olive flounder (Paralichthys olivaceus)
ISSN号	0006-3088
DOI	10.2478/s11756-020-00677-7
通讯作者	Tan, Xungang(tanx@qdio.ac.cn) ; You, Feng(youfeng@qdio.ac.cn)
英文摘要	As a new breeding technology, genome editing becomes a powerful tool owing to its high efficiency of gene targeting. In CRISPR/Cas9 system, how to efficiently transfer gRNA and Cas9 mRNA into embryos is an important step. Though microinjection is the most common method for operating on fish embryos, it is not easy to inject the RNA into pelagic and telolecithal eggs with hard egg chorion, such as the olive flounder (Paralichthys olivaceus) eggs. Therefore, an efficient and simple technology is urgently needed for this kind of study. In the present study, we used the electroporation method to introduce foreign gene into the flounder eggs. The results showed that the proper electroporation condition was 3 pulses for 1 millisecond (ms), 50 ms interval, at 25 V with high survival rate. Under this condition, the effect of CRISPR/Cas9 system on genome editing by using two different genes, myomaker and gonadal soma derived factor (gsdf) was investigated. Around 12% and 7% of the electroporated embryos for myomaker and gsdf hatched, respectively. The mutation sites including insert and deletion mutations at the candidate sites were visible for both targeted genes in the hatched larvae. The checked frame-shift and start codon deletion mutations would lead to complete destruction of these genes' structure. Above results implied that CRISPR/Cas9 system could work well in marine fish with pelagic eggs by using electroporation, and genome editing could be achieved on a large scale which may be useful for study of gene function in marine fish.
资助项目	National Key R&D Program of China[2018YFD0900202] ; National Sciences of Foundation of China[31672636] ; National Sciences of Foundation of China[31772834]
WOS研究方向	Life Sciences & Biomedicine - Other Topics
语种	英语
出版者	SPRINGER
WOS记录号	WOS:000609070200004
内容类型	期刊论文
源URL	[http://ir.qdio.ac.cn/handle/337002/170063]
专题	海洋研究所_实验海洋生物学重点实验室
通讯作者	Tan, Xungang; You, Feng
作者单位	1.Chinese Acad Sci, Ctr Ocean Megasci, Inst Oceanol, CAS Key Lab Expt Marine Biol, 7 Nanhai Rd, Qingdao 266071, Peoples R China 2.Pilot Natl Lab Marine Sci & Technol Qingdao, Lab Marine Biol & Biotechnol, Qingdao 266237, Peoples R China 3.Shenghang Aquat Sci & Technol Co Ltd, Weihai, Peoples R China 4.Univ Chinese Acad Sci, Beijing 100049, Peoples R China 5.Weihai Fisheries Technol Promot Stn, Weihai, Peoples R China
推荐引用方式 GB/T 7714	Wang, Ling,Tan, Xungang,Wu, Zhihao,et al. Targeted mutagenesis in the olive flounder (Paralichthys olivaceus) using the CRISPR/Cas9 system with electroporation[J]. BIOLOGIA,2021:8.
APA	Wang, Ling.,Tan, Xungang.,Wu, Zhihao.,Wang, Lijuan.,Jiao, Shuang.,...&You, Feng.(2021).Targeted mutagenesis in the olive flounder (Paralichthys olivaceus) using the CRISPR/Cas9 system with electroporation.BIOLOGIA,8.
MLA	Wang, Ling,et al."Targeted mutagenesis in the olive flounder (Paralichthys olivaceus) using the CRISPR/Cas9 system with electroporation".BIOLOGIA (2021):8.