Simple promotion of Cas9 and Cas12a expression improves gene targeting via an all-in-one strategy

文献类型： 外文期刊

第一作者： Cheng, Yiqiu

作者： Cheng, Yiqiu;Zhang, Lei;Li, Jing;Dang, Xiaofei;Miki, Daisuke;Cheng, Yiqiu;Zhang, Lei;Li, Jing;Zhu, Jian-Kang;Zhu, Jian-Kang;Zhu, Jian-Kang;Shimada, Hiroaki

作者机构： Chinese Acad Sci, Shanghai Ctr Plant Stress Biol, CAS Ctr Excellence Mol Plant Sci, Shanghai, Peoples R China;Univ Chinese Acad Sci, Beijing, Peoples R China;Southern Univ Sci & Technol, Inst Adv Biotechnol, Shenzhen, Peoples R China;Southern Univ Sci & Technol, Sch Life Sci, Shenzhen, Peoples R China;Chinese Acad Agr Sci, Ctr Adv Bioind Technol, Beijing, Peoples R China;Tokyo Univ Sci, Dept Biol Sci & Technol, Tokyo, Japan

关键词： genome engineering; CRISPR/Cas9; CRISPR/ttCas12a; RPS5a; all-in-one; gene targeting (GT); transcriptional and translational enhancers

期刊名称：FRONTIERS IN PLANT SCIENCE （ 2022影响因子：5.6； 五年影响因子：6.8 ）

ISSN： 1664-462X

年卷期： 2024 年 15 卷

页码：

收录情况： SCI

摘要： Gene targeting (GT) is a promising tool for precise manipulation of genome sequences, however, GT in seed plants remains a challenging task. The simple and direct way to improve the efficiency of GT via homology-directed repair (HDR) is to increase the frequency of double-strand breaks (DSBs) at target sites in plants. Here we report an all-in-one approach of GT in Arabidopsis by combining a transcriptional and a translational enhancer for the Cas expression. We find that facilitating the expression of Cas9 and Cas12a variant by using enhancers can improve DSB and subsequent knock-in efficiency in the Arabidopsis genome. These results indicate that simply increasing Cas protein expression at specific timings - egg cells and early embryos - can improve the establishment of heritable GTs. This simple approach allows for routine genome engineering in plants.

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