CABE-RY: A PAM-flexible dual-mutation base editor for reliable modeling of multi-nucleotide variants

Wanyu Tao; Qing Liu; Shisheng Huang; Xin Wang; Shiyuan Qu; Junfan Guo; Danfeng Ou; Guanglei Li; Yu Zhang; Xiangmin Xu; Xingxu Huang

doi:10.1016/j.omtn.2021.07.016

CABE-RY: A PAM-flexible dual-mutation base editor for reliable modeling of multi-nucleotide variants

Mol Ther Nucleic Acids. 2021 Jul 29:26:114-121. doi: 10.1016/j.omtn.2021.07.016. eCollection 2021 Dec 3.

Authors

Wanyu Tao^{1

2}, Qing Liu³, Shisheng Huang^{1

2}, Xin Wang^{1

2}, Shiyuan Qu^{1

2}, Junfan Guo^{1

2}, Danfeng Ou⁴, Guanglei Li^{1

2}, Yu Zhang^{1

2}, Xiangmin Xu^{3

5}, Xingxu Huang^{1

2}

Affiliations

¹ School of Life Science and Technology, ShanghaiTech University, Shanghai 201210, China.
² University of Chinese Academy of Sciences, Beijing 100049, China.
³ Department of Medical Genetics, School of Basic Medical Sciences, Southern Medical University, Guangzhou, Guangdong 510515, China.
⁴ International Academy of Optoelectronics at Zhaoqing, South China Normal University, Zhaoqing 526238, China.
⁵ Guangdong Genetic Testing Engineering Research Center, Guangzhou, Guangdong 510515, China.

Abstract

Multi-nucleotide variants (MNVs) represent an important type of genetic variation and have biological and clinical significance. To simulate MNVs, we designed four dual-mutation base editors combining hA3A(Y130F), TadA8e(V106W), and protospacer adjacent motif (PAM)-flexible SpRY and selected cytosine and adenine base editor-SpRY (CABE-RY), which had the best editing performance, for further study. Characterization and comparison showed that CABE-RY had a smaller DNA editing window and lower RNA off-target edits than the corresponding single base editors. Thus, we have established a versatile tool to efficiently simulate MNVs over the genome, which could be very useful for functional studies on MNVs in humans.

Keywords: CRISPR; MNVs; PAM-flexible; dual-mutation base editor; simultaneous A/C conversions.