胞嘧啶脱氨酶
Cas9
胞苷脱氨酶
活化诱导(胞苷)脱氨酶
清脆的
DNA
胞嘧啶
生物
基因组编辑
计算生物学
分子生物学
遗传学
基因
遗传增强
体细胞突变
抗体
B细胞
作者
Jizeng Zhou,Yang Liu,Yuhui Wei,Shuwen Zheng,Shixue Gou,Tao Chen,Yang Yang,Ting Lan,Min Chen,Yuan Liao,Quanjun Zhang,Chengcheng Tang,Yu Liu,Yunqin Wu,Xiaohua Peng,Minghui Gao,Junwei Wang,Kun Zhang,Liangxue Lai,Qingjian Zou
标识
DOI:10.1016/j.ymthe.2022.04.010
摘要
Predictable DNA off-target effect is one of the major safety concerns for the application of cytosine base editors (CBEs). To eliminate Cas9-dependent DNA off-target effects, we designed a novel effective CBE system with dual guiders by combining CRISPR with transcription activator-like effector (TALE). In this system, Cas9 nickase (nCas9) and cytosine deaminase are guided to the same target site to conduct base editing by single-guide RNA (sgRNA) and TALE, respectively. However, if nCas9 is guided to a wrong site by sgRNA, it will not generate base editing due to the absence of deaminase. Similarly, when deaminase is guided to a wrong site by TALE, base editing will not occur due to the absence of single-stranded DNA. In this way, Cas9- and TALE-dependent DNA off-target effects could be completely eliminated. Furthermore, by fusing TALE with YE1, a cytidine deaminase with minimal Cas9-independent off-target effect, we established a novel CBE that could induce efficient C-to-T conversion without detectable Cas9- or TALE-dependent DNA off-target mutations.
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