摘要由规律成簇间隔短回文重复序列(clustered regularly interspaced short palindromic repeats，CRISPR)以及CRISPR相关蛋白９(CRISPR-associated protein 9, Cas9)组成的系统被发现广泛存在于细菌及古菌中，是机体长期进化形成的由RNA指导的降解外源遗传物质的适应性免疫系统。由于该系统可以识别靶向序列完成DNA的双链切割，因此从2013年起，CRISPR/Cas9系统即被改造为基因编辑工具。作为一种新型的基因组编辑技术，CRISPR/Cas9系统具有设计简单、特异性强、效率高等优点，为基因组定向改造调控和应用带来了突破性的革命，并且迅速在基础理论、转基因动物生产、基因诊断和临床治疗等领域中得到广泛的研究与应用。然而，CRISPR/Cas9也存在着脱靶效应和外源基因插入困难等一些亟待解决的问题，这在一定程度上限制了CRISPR/Cas9 的应用。因此，近年来围绕CRISPR系统改进获得了一系列可用于基因编辑与基因表达修饰的新工具。本文介绍了CRISPR系统的组成、工作原理，并综述了近几年来基于CRISPR系统开发的几种新型基因编辑工具以及基因表达修饰工具的研究进展和应用，并对其应用前景和发展方向进行了展望。
Abstract：Bacteria and archaea have evolved an adaptive immune system, which was known as clustered regularly interspaced short palindromic repeats (CRISPR) / CRISPR -associated protein (Cas 9) system, and that uses short RNA to degrade the target sequences present in invading viral and phage DNAs. This system can identify the target sequence and cut the double-stranded DNA. So, it has been harnessed by thousands of laboratories for genome editing applications in a variety of experimental model systems since 2013. As a new type of genome editing techniques, the CRISPR/Cas9 system has advantages in simple in design, strong specificity and high efficiency. CRISPR/Cas9 system has brought a breakthrough in genome directional regulation and application, and rapidly applied in the basic theory, transgenic animal production, genetic diagnosis, clinical therapy fields. Whereas, every new technology has its limitations, CRISPR/Cas9 system also exists off-target effects and difficulty to insert genes. This limits its applications to some extent. Therefore, around the CRISPR system, many scientists improved a series of modification that can be used for the gene editing and gene expression of new tools in recent years. Here, we reviewed the research progress and application of new gene editing tools and gene expression modification based on the CRISPR system evolution and looking forward to the application prospect and development direction.
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