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Optimized guide RNA structure for genome editing via Cas9

2017; Impact Journals LLC; Volume: 8; Issue: 55 Linguagem: Inglês

10.18632/oncotarget.21607

1949-2553

Jianyong Xu, Wei Lian, Yuning Jia, Lingyun Li, Zhong Huang,

RNA and protein synthesis mechanisms

// Jianyong Xu 1, 2 , Wei Lian 1, 2 , Yuning Jia 1, 2 , Lingyun Li 1, 2 and Zhong Huang 1, 2 1 Institute of Biological Therapy, Shenzhen University, Shenzhen, P.R. China 2 Department of Pathogen Biology and Immunology, School of Medicine, Shenzhen University, Shenzhen, P.R. China Correspondence to: Jianyong Xu, email: xujianyong@szu.edu.cn Zhong Huang, email: zhuang809@126.com Keywords: RNA guided endonuclease, genome editing, Cas9, guide RNA, gRNA Received: August 09, 2017 Accepted: August 28, 2017 Published: October 07, 2017 ABSTRACT The genome editing tool Cas9-gRNA (guide RNA) has been successfully applied in different cell types and organisms with high efficiency. However, more efforts need to be made to enhance both efficiency and specificity. In the current study, we optimized the guide RNA structure of Streptococcus pyogenes CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats)/Cas (CRISPR-associated) system to improve its genome editing efficiency. Comparing with the original functional structure of guide RNA, which is composed of crRNA and tracrRNA, the widely used chimeric gRNA has shorter crRNA and tracrRNA sequence. The deleted RNA sequence could form extra loop structure, which might enhance the stability of the guide RNA structure and subsequently the genome editing efficiency. Thus the genome editing efficiency of different forms of guide RNA was tested. And we found that the chimeric structure of gRNA with original full length of crRNA and tracrRNA showed higher genome editing efficiency than the conventional chimeric structure or other types of gRNA we tested. Therefore our data here uncovered the new type of gRNA structure with higher genome editing efficiency.