346 0

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
Files in This Item:
There are no files associated with this item.
Export
RIS (EndNote)
XLS (Excel)
XML


qrcode

Items in DSpace are protected by copyright, with all rights reserved, unless otherwise indicated.

BROWSE