Wei Chunjie, Liu Hao, Wang Wenwen, Luo Pengyu, Chen Qiuling, Li Rou, Wang Chong, Botella José Ramón, Zhang Hui
Shanghai Key Laboratory of Plant Molecular Sciences, Development Center of Plant Germplasm Resources, College of Life Sciences, Shanghai Normal University, Shanghai, China.
School of Agriculture and Food Sciences, University of Queensland, Brisbane, QLD, Australia.
Front Plant Sci. 2022 Apr 25;13:865848. doi: 10.3389/fpls.2022.865848. eCollection 2022.
Recently developed base editors provide a powerful tool for plant research and crop improvement. Although a number of different deaminases and Cas proteins have been used to improve base editors the editing efficiency, and editing window are still not optimal. Fusion of a non-sequence-specific single-stranded DNA-binding domain (DBD) from the human Rad51 protein between Cas9 nickase and the deaminase has been reported to dramatically increase the editing efficiency and expand the editing window of base editors in the mammalian cell lines and mouse embryos. We report the use of this strategy in rice, by fusing a rice codon-optimized human Rad51 DBD to the cytidine base editors AncBE4max, AncBE4max-NG, and evoFERNY. Our results show that the addition of Rad51 DBD did not increase editing efficiency in the major editing window but the editing range was expanded in all the three systems. Replacing the human Rad51 DBD with the rice Rad51 DBD homolog also expanded the editing window effectively.
最近开发的碱基编辑器为植物研究和作物改良提供了一个强大的工具。尽管已经使用了多种不同的脱氨酶和Cas蛋白来改进碱基编辑器，但编辑效率和编辑窗口仍不理想。据报道，在Cas9切口酶和脱氨酶之间融合来自人类Rad51蛋白的非序列特异性单链DNA结合结构域（DBD），可显著提高哺乳动物细胞系和小鼠胚胎中碱基编辑器的编辑效率并扩大其编辑窗口。我们报告了该策略在水稻中的应用，即将水稻密码子优化的人类Rad51 DBD与胞嘧啶碱基编辑器AncBE4max、AncBE4max-NG和evoFERNY融合。我们的结果表明，添加Rad51 DBD并未提高主要编辑窗口中的编辑效率，但在所有三个系统中编辑范围均得到了扩大。用水稻Rad51 DBD同源物替代人类Rad51 DBD也有效地扩大了编辑窗口。
