A Cas-embedding strategy for minimizing off-target effects of DNA base editors
Yajing Liu, Changyang Zhou, Shisheng Huang, Lu Dang, Wei Yu, Jun He, Yingsi Zhou, Shaoshuai Mao, Wanyu Tao, Yu Zhang, Hui Yang, Xingxu Huang, Tian Chi
Abstract
DNA base editors, typically comprising editing enzymes fused to the N-terminus of nCas9, display off-target effects on DNA and/or RNA, which have remained an obstacle to their clinical applications. Off-target edits are typically countered via rationally designed point mutations, but the approach is tedious and not always effective. Here, we report that the off-target effects of both A > G and C > T editors can be dramatically reduced without compromising the on-target editing simply by inserting the editing enzymes into the middle of nCas9 at tolerant sites identified using a transposon-based genetic screen. Furthermore, employing this Cas-embedding strategy, we have created a highly specific editor capable of efficient C > T editing at methylated and GC-rich sequences.