Precise large-fragment deletions in mammalian cells and mice generated by dCas9-controlled CRISPR/Cas3
Jinze Li, Ding Zhao, Tao Zhang, Haoyang Xiong, Mingyang Hu, Hongmei Liu, Feiyu Zhao, Xiaodi Sun, Peng Fan, Yuqiang Qian, Di Wang, Liangxue Lai, Tingting Sui, Zhanjun Li
Abstract
Currently, the Cas9 and Cas12a systems are widely used for genome editing, but their ability to precisely generate large chromosome fragment deletions is limited. Type I-E CRISPR mediates broad and unidirectional DNA degradation, but controlling the size of Cas3-mediated DNA deletions has proven elusive thus far. Here, we demonstrate that the endonuclease deactivation of Cas9 (dCas9) can precisely control Cas3-mediated large-fragment deletions in mammalian cells. In addition, we report the elimination of the Y chromosome and precise retention of the Sry gene in mice using CRISPR/Cas3 and dCas9-controlled CRISPR/Cas3, respectively. In conclusion, dCas9-controlled CRISPR/Cas3-mediated precise large-fragment deletion provides an approach for establishing animal models by chromosome elimination. This method also holds promise as a potential therapeutic strategy for treating fragment mutations or human aneuploidy diseases that involve additional chromosomes.