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Kim, Dae Sik
유전체 교정 연구단
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Genome-wide specificity of dCpf1 cytidine base editors

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Title
Genome-wide specificity of dCpf1 cytidine base editors
Author(s)
Daesik Kim; Kayeong Lim; Da-eun Kim; Jin-Soo Kim
Subject
DNA
Publication Date
2020-08
Journal
NATURE COMMUNICATIONS, v.11, no.1, pp.4072
Publisher
NATURE PUBLISHING GROUP
Abstract
© 2020 The Author(s). Cpf1-linked base editors broaden the targeting scope of programmable cytidine deaminases by recognizing thymidine-rich protospacer-adjacent motifs (PAM) without inducing DNA double-strand breaks (DSBs). Here we present an unbiased in vitro method for identifying genome-wide off-target sites of Cpf1 base editors via whole genome sequencing. First, we treat human genomic DNA with dLbCpf1-BE ribonucleoprotein (RNP) complexes, which convert C-to-U at on-target and off-target sites and, then, with a mixture of E. coli uracil DNA glycosylase (UDG) and DNA glycosylase-lyase Endonuclease VIII, which removes uracil and produces single-strand breaks (SSBs) in vitro. Whole-genome sequencing of the resulting digested genome (Digenome-seq) reveals that, on average, dLbCpf1-BE induces 12 SSBs in vitro per crRNA in the human genome. Off-target sites with an editing frequency as low as 0.1% are successfully identified by this modified Digenome-seq method, demonstrating its high sensitivity. dLbCpf1-BEs and LbCpf1 nucleases often recognize different off-target sites, calling for independent analysis of each tool
URI
https://pr.ibs.re.kr/handle/8788114/7706
DOI
10.1038/s41467-020-17889-9
ISSN
2041-1723
Appears in Collections:
Center for Genome Engineering(유전체 교정 연구단) > 1. Journal Papers (저널논문)
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