Digenome-seq: genome-wide profiling of CRISPR-Cas9 off-target effects in human cells
- Title
- Digenome-seq: genome-wide profiling of CRISPR-Cas9 off-target effects in human cells
- Author
- 배상수
- Keywords
- ZINC-FINGER NUCLEASES; CRISPR/CAS9 SYSTEMS; GUIDE RNA; CAS9; SPECIFICITY; ENDONUCLEASE; DNA; RIBONUCLEOPROTEINS; MUTATIONS; NICKASES; DNA sequencing; Genetic engineering; Genomics
- Issue Date
- 2015-03
- Publisher
- NATURE PUBLISHING GROUP
- Citation
- NATURE METHODS, v. 12, NO 3, Page. 237-243
- 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 5' 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
- http://www.nature.com/nmeth/journal/v12/n3/abs/nmeth.3284.htmlhttp://hdl.handle.net/20.500.11754/22846
- ISSN
- 1548-7091; 1548-7105
- DOI
- 10.1038/NMETH.3284
- Appears in Collections:
- COLLEGE OF NATURAL SCIENCES[S](자연과학대학) > CHEMISTRY(화학과) > Articles
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