As versatile and robust genome editing tools, clustered regularly interspaced short palindromic repeats (CRISPR) technologies have been broadly used in basic research, biotechnology, and therapeutic development. Off-target mutagenesis by CRISPR systems has been demonstrated, and various methods have been developed to markedly increase their specificity. In this review, we highlight the efforts of producing and modifying guide RNA (gRNA) to minimize off-target activities, including sequence and structure design, tuning expression and chemical modification. The modalities of gRNA engineering can be applied across CRISPR systems. In conjunction with CRISPR protein effectors, the engineered gRNA enables efficient and precise genome editing.

作为多功能且强大的基因组编辑工具，成簇规则间隔短回文重复（CRISPR）技术已广泛应用于基础研究、生物技术和治疗开发。 CRISPR 系统的脱靶诱变已得到证实，并且已开发出多种方法来显着提高其特异性。在这篇综述中，我们重点介绍了生产和修饰指导 RNA (gRNA) 以尽量减少脱靶活动的努力，包括序列和结构设计、调整表达和化学修饰。 gRNA 工程的模式可以应用于整个 CRISPR 系统。与 CRISPR 蛋白效应器结合，工程 gRNA 能够实现高效、精确的基因组编辑。