数字农科院2.0

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

文献类型： 外文期刊

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

作者机构：

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

期刊名称： Frontiers in Plant Science

ISSN： 1664-462X

年卷期： 2024 年 15 卷

页码：

收录情况： SCIE（2024版）

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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