CRISPR/Cas9的应用及脱靶效应研究进展

Progress of application and off-target effects of CRISPR/Cas9

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,sg RNA)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sg RNA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。

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.