| Literature DB >> 29637228 |
Han Wu1,2,3, Qishuai Liu1,2,3, Hui Shi1,2,3, Jingke Xie1,2,3, Quanjun Zhang1,3, Zhen Ouyang1,3, Nan Li1,2,3, Yi Yang4, Zhaoming Liu1,3, Yu Zhao1,3, Chengdan Lai1,3, Degong Ruan1,2,3, Jiangyun Peng1,2,3, Weikai Ge1,2,3, Fangbing Chen1,2,3, Nana Fan1,3, Qin Jin1,2,3, Yanhui Liang1,2,3, Ting Lan1,2,3, Xiaoyu Yang1,5, Xiaoshan Wang1,2,3, Zhiyong Lei6,7, Pieter A Doevendans6,7, Joost P G Sluijter6,7, Kepin Wang8,9, Xiaoping Li10,11, Liangxue Lai12,13,14.
Abstract
CRISPR/Cpf1 features a number of properties that are distinct from CRISPR/Cas9 and provides an excellent alternative to Cas9 for genome editing. To date, genome engineering by CRISPR/Cpf1 has been reported only in human cells and mouse embryos of mammalian systems and its efficiency is ultimately lower than that of Cas9 proteins from Streptococcus pyogenes. The application of CRISPR/Cpf1 for targeted mutagenesis in other animal models has not been successfully verified. In this study, we designed and optimized a guide RNA (gRNA) transcription system by inserting a transfer RNA precursor (pre-tRNA) sequence downstream of the gRNA for Cpf1, protecting gRNA from immediate digestion by 3'-to-5' exonucleases. Using this new gRNAtRNA system, genome editing, including indels, large fragment deletion and precise point mutation, was induced in mammalian systems, showing significantly higher efficiency than the original Cpf1-gRNA system. With this system, gene-modified rabbits and pigs were generated by embryo injection or somatic cell nuclear transfer (SCNT) with an efficiency comparable to that of the Cas9 gRNA system. These results demonstrated that this refined gRNAtRNA system can boost the targeting capability of CRISPR/Cpf1 toolkits.Entities:
Keywords: CRISPR/Cpf1; Genome editing; Pig; Rabbit; gRNAtRNA system
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Year: 2018 PMID： 29637228 DOI： 10.1007/s00018-018-2810-3
Source DB: PubMed Journal: Cell Mol Life Sci ISSN： 1420-682X Impact factor: 9.261