Improved design and analysis of CRISPR knockout screens

被引：33

作者：

Chen, Chen-Hao ^{[1
,2
,3
,4
]}

Xiao, Tengfei ^{[1
,5
,6
]}

Xu, Han ^{[1
,2
,3
,7
]}

Jiang, Peng ^{[1
,2
,3
]}

Meyer, Clifford A. ^{[1
,2
,3
]}

Li, Wei ^{[1
,2
,3
,8
,9
]}

Brown, Myles ^{[1
,5
,6
]}

Liu, X. Shirley ^{[1
,2
,3
]}

机构：

[1] Dana Farber Canc Inst, Ctr Funct Canc Epigenet, Boston, MA 02115 USA

[2] Dana Farber Canc Inst, Dept Biostat & Computat Biol, Boston, MA 02115 USA

[3] Harvard Sch Publ Hlth, Boston, MA 02115 USA

[4] Harvard Med Sch, Biol & Biomed Sci Program, Boston, MA 02115 USA

[5] Dana Farber Canc Inst, Dept Med Oncol, Boston, MA 02115 USA

[6] Harvard Med Sch, Boston, MA 02115 USA

[7] Univ Texas MD Anderson Canc Ctr, Dept Epigenet & Mol Carcinogenesis, Smithville, TX 78957 USA

[8] Childrens Natl Hlth Syst, Ctr Genet Med Res, Washington, DC 20010 USA

[9] George Washington Sch Med & Hlth Sci, Dept Genom & Precis Med, Washington, DC 20010 USA

来源：

BIOINFORMATICS | 2018年 / 34卷 / 23期

基金：

美国国家卫生研究院;

关键词：

OFF-TARGET; FUNCTIONAL GENOMICS; GENE ESSENTIALITY; SPECIFICITY; INTEGRATION; IDENTIFICATION; CELLS;

D O I：

10.1093/bioinformatics/bty450

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

Motivation: Genome-wide clustered, regularly interspaced, short palindromic repeat (CRISPR)-Cas9 screen has been widely used to interrogate gene functions. However, the rules to design better libraries beg further refinement. Results: We found single guide RNA (sgRNA) outliers are characterized by higher G-nucleotide counts, especially in regions distal from the PAM motif and are associated with stronger off-target activities. Furthermore, using non-targeting sgRNAs as negative controls lead to strong bias, which can be mitigated by using sgRNAs targeting multiple 'safe harbor' regions. Custom-designed screens confirmed our findings and further revealed that 19 nt sgRNAs consistently gave the best signal-to-noise ratio. Collectively, our analysis motivated the design of a new genome-wide CRISPR/Cas9 screen library and uncovered some intriguing properties of the CRISPR-Cas9 system.

引用

页码：4095 / 4101

页数：7

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