Digenome-seq: Genome-wide profiling of CRISPR-Cas9 off-target effects in human cells Highly Cited Paper

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Title
Digenome-seq: Genome-wide profiling of CRISPR-Cas9 off-target effects in human cells
Author(s)
Daesik Kim; Sangsu Bae; Jeongbin Park; Eunji Kim; Seokjoong Kim; Hye Ryeong Yu; Jinha Hwang; Jong-Il Kim; Jin-Soo Kim
Publication Date
2015-03
Journal
NATURE METHODS, v.12, no.3, pp.237 - 243
Publisher
NATURE PUBLISHING GROUP
Abstract
Although RNA-guided genome editing via the CRISPR-Cas9 system is now widely used in biomedical research, genome-wide target specificities of Cas9 nucleases remain controversial. Here we present Digenome-seq, in vitro Cas9-digested whole-genome sequencing, to profile genome-wide Cas9 off-target effects in human cells. This in vitro digest yields sequence reads with the same 5â €2 ends at cleavage sites that can be computationally identified. We validated off-target sites at which insertions or deletions were induced with frequencies below 0.1%, near the detection limit of targeted deep sequencing. We also showed that Cas9 nucleases can be highly specific, inducing off-target mutations at merely several, rather than thousands of, sites in the entire genome and that Cas9 off-target effects can be avoided by replacing 'promiscuous' single guide RNAs (sgRNAs) with modified sgRNAs. Digenome-seq is a robust, sensitive, unbiased and cost-effective method for profiling genome-wide off-target effects of programmable nucleases including Cas9
URI
https://pr.ibs.re.kr/handle/8788114/1608
ISSN
1548-7091
Appears in Collections:
Center for Genome Engineering(유전체 교정 연구단) > Journal Papers (저널논문)
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