动物基因组编辑中提升CRISPR/Cas9介导的同源重组效率研究进展

Research Advancement of CRISPR/Cas9 Directed Homologous Recombination Efficiency Improvements in Mammal Genome Editing

基因组编辑技术可以对DNA或RNA进行精准改造,极大地促进了生命科学的发展。CRISPR/Cas9系统在靶位点诱导DNA发生双链或单链损伤,细胞对损伤部位采用无供体模板的非同源末端连接(non-homologous end joining, NHEJ)或有供体模板的同源重组(homologous recombination, HR)修复。基于HR的基因组编辑策略通常被用于获得DNA的精准改造,而NHEJ在动物DNA损伤修复中起主导作用。为了提升HR效率,研究人员设计了多种方案,包括CRISPR/Cas9系统优化和DNA修复通路调控等。从DNA损伤修复途径、Cas9变体选择、sgRNA设计、供体模板设计、DNA修复途径相关蛋白功能调控、供体模板募集效率提升、细胞周期调控及编辑细胞生存效率提升等方面详细综述了相关研究成果,发现尚未开发出放之四海而皆准的HR提升策略,基于HR的基因组编辑需要针对具体案例制定个体化策略。旨在为动物基因组编辑中提升CRISPR/Cas9介导的HR效率研究提供理论参考,为动物基因功能分析、基因治疗和经济动物基因编辑育种提供帮助。

The genome editing system can bring the precise modification of DNA or RNA, which provides great help for the development of life sciences. The CRISPR/Cas9 could induce double or single strand DNA damage at target sites. The damages are repaired by non-homologous end joining(NHEJ) without donor template or homologous recombination(HR) with donor template in cells. The HR-based genome editing strategy is common to generate precise modification of DNA. However, NHEJ plays a dominant role in mammal DNA repair. To improve the HR efficiency, the researchers designed multiple strategies, which include genome editing system optimization and DNA repair pathway regulation. The related research achievements from the aspects of DNA damage repair pathway, Cas9 mutation selection, sgRNA design, donor template design, functional regulation of DNA repair pathway related proteins, improvement of donor template recruitment efficiency, cell cycle regulation and improvement of editing cell survival efficiency were reviewed. A one-size-fits-all HR promotion strategy has not yet been developed. The case-specific strategies of HR-based are required during relative research. This review can provide theoretical reference for improving the efficiency of CRISPR/Cas9-mediated HR in animal genome editing and provide help for animal gene function analysis, gene therapy and economic animal breeding through gene editing.