文献类型：期刊文献

中文题名：CRISPR/Cas系统中工程化gRNA技术的研究和应用

英文题名：Research and Application of the Engineered gRNA Technology in the CRISPR/Cas System

作者：谈鎏[1];叶邦策[2];尹斌成[2]

机构：[1]浙江工业大学药学院,长三角绿色制药协同创新中心,杭州310014;[2]华东理工大学生物反应器工程国家重点实验室,上海200237

年份：2024

卷号：40

期号：8

起止页码：1078

中文期刊名：中国生物化学与分子生物学报

外文期刊名：Chinese Journal of Biochemistry and Molecular Biology

收录：CSTPCD;;Scopus;北大核心:【北大核心2023】；CSCD:【CSCD_E2023_2024】；PubMed;

基金：国家自然科学基金(No.22374047,No.22134003)资助。

语种：中文

中文关键词：CRISPR/Cas系统;gRNA工程化;基因编辑;特异性

外文关键词：clustered regularly interspaced short palindromic repeats(CRISPR)/CRISPR-associated(Cas)system;gRNA engineering;genome editing;specificity

摘要：CRISPR/Cas是原核生物在进化过程中获得的一种免疫防御系统,用于抵抗外来遗传物质的入侵,近年来被开发成为高效的基因编辑、基因调控以及分子诊断工具。其可编程靶向机制揭开了利用该系统进行基因组操作的序幕,并允许在活性范围内实现动态调节和控制基因表达。作为现有基因修饰手段中灵活性最强和成本最低的技术之一,已被广泛应用于临床疾病治疗、工农业生产、可持续染料开发和化学品加工等领域。随着对CRISPR/Cas系统的不断深入挖掘和探索,大量研究报道了gRNA工程改造及优化方法,包括改变间隔序列长度、调节恒定区和可变区的结构、向末端或中间延伸添加额外功能序列及化学合成修饰等,以期降低脱靶突变率,提高作用效率,充分激发CRISPR基因操纵工具在生物医学方面的潜力。基于此,本综述将介绍CRISPR/Cas9和CRISPR/Cas12系统中gRNA工程化设计方法及应用研究的最新进展,分析探讨了当前工程化gRNA技术面临的机遇和挑战,旨在为获得性能更加优异的gRNA提供思路和方向,从而提高利用CRISPR/Cas系统探测人类细胞基因组的能力,进一步为可编程生物学带来更多可能性。
The CRISPR/Cas is an immune defense system acquired by prokaryotes to resist the invasion of foreign genetic materials during their evolutionary process.In recent years,it has been developed into an efficient tool for genome editing,gene regulation and molecular diagnosis.Its programmable targeting mechanism has opened the door to use this system for genome manipulation and allows for dynamic regulation and control of gene expression within its activity range.As one of the most flexible and cost-effective techniques among existing gene modification methods,it has been widely applied in clinical disease treatment,industrial and agricultural production,sustainable dye development,chemical processing and many other fields.With the continuous investigation and exploration of the CRISPR/Cas system,a large number of studies have been reported on the engineering modification and optimization approaches of gRNA,including changing the length of the spacer region,regulating the structure of constant and variable sequence parts,adding extra functional sequences through the end or middle extension,and chemical synthesis modifications,in order to reduce off-target and mutation rates,improve the efficiency of the CRISPR system,and fully stimulate the potential of CRISPR gene manipulation tools in biomedical fields.Based on this,this review will introduce the latest progress in gRNA engineering design strategies and application research of CRISPR/Cas9 and CRISPR/Cas12 systems,analyze and discuss the opportunities and challenges in the current gRNA engineering technology,aiming to provide ideas and reference directions for obtaining gRNAs with better performance,thereby effectively improving the ability to probe the human genomes using the CRISPR/Cas system and bringing more possibilities to programmable biology.