CRISPRCas9的应用及脱靶效应研究进展.docxVIP

Hereditas (Beijing) 2015 年 10 月 , 37(10): 1003―1010 综 述 CRISPR/Cas9 的应用及脱靶效应研究进展 郑武，谷峰 温州医科大学附属眼视光医院，眼视光学和视觉科学国家重点实验室培育基地，卫生部视觉科学研究重点实验室，温州 325027 摘要: CRISPR/Cas9 基因编辑技术在生命科学领域掀起了一场全新的技术革命，该技术可以对基因组特定位点进行靶向编辑，包括缺失、插入、修复等。CRISPR/Cas9 比锌指核酸酶 (ZFNs)和转录激活因子样效应物核酸酶(TALENs)技术更易于操作，而且更高效。CRISPR/Cas9 系统中的向导 RNA(Single guide RNA, sgRNA)是一段与目标 DNA 片段匹配的 RNA 序列，指导 Cas9 蛋白对基因组进行识别。研究发现，设计的 sgRNA 会与非靶点 DNA 序列错配，引入非预期的基因突变，即脱靶效应(Off-target effects)。脱靶效应严重制约了 CRISPR/Cas9 基因编辑技术的广泛应用。为了避免脱靶效应，研究者对影响脱靶效应的因素进行了系统研究并提出了许多降低脱靶效应的方法。文章总结了 CRISPR/Cas9 系统的应用及脱靶效应研究进展，以期为相关领域的工作提供参考。 关键词: 基因组编辑；CRISPR/Cas9；脱靶效应；向导 RNA Progress of application and off-target effects of CRISPR/Cas9 Wu Zheng, Feng Gu State Key Laboratory Cultivating Base of Optometry and Visual Science, Key Laboratory of Vision Science, Ministry of Health, The Eye Hospital of Wenzhou Medical University, Wenzhou 325027 China Abstract: The clustered regulatory interspaced short palindromic repeat/Cas9 (CRISPR/Cas9) system mediates genome editing and is revolutionizing genetic researches. Scientists are able to manipulate the gene of interest from any organism with CRISPR/Cas9. Compared with zinc finger nucleases (ZFNs) and transcription activator-like ef- fector nucleases (TALENs) technologies, the CRISPR/Cas9 technology provides an easy and efficient approach to manipulate the genome. In this system, sgRNA (Single guide RNA), a short RNA matching the targeted DNA frag- ment, guides the CRISPR/Cas9 to interrogate the genome. Because sgRNA can tolerate certain mismatches to the DNA targets and thereby promote undesired off-target mutagenesis, the key limit of this technology is off-target ef- fects. To eliminate the off-target effects, different strategies have been adopted. In this review, we summarize the application of CRISPR/Cas9 and different strategies for addressing off-target effects. Keywords: genome editing; CRISPR/Cas9; off-target effects; sgRNA 收稿日期: 2015?02?09; 修