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Nat Commun. 2017 May 23;8:15403. doi: 10.1038/ncomms15403.

Complementary information derived from CRISPR Cas9 mediated gene deletion and suppression.

Author information

1
Broad Institute of Harvard and MIT, 415 Main Street, Cambridge, Massachusetts 02142, USA.
2
Department of Medical Oncology, Dana-Farber Cancer Institute, Harvard Medical School, 450 Brookline Avenue, Boston, Massachusetts 02215, USA.

Abstract

CRISPR-Cas9 provides the means to perform genome editing and facilitates loss-of-function screens. However, we and others demonstrated that expression of the Cas9 endonuclease induces a gene-independent response that correlates with the number of target sequences in the genome. An alternative approach to suppressing gene expression is to block transcription using a catalytically inactive Cas9 (dCas9). Here we directly compare genome editing by CRISPR-Cas9 (cutting, CRISPRc) and gene suppression using KRAB-dCas9 (CRISPRi) in loss-of-function screens to identify cell essential genes. CRISPRc identified 98% of previously defined cell essential genes. After optimizing library construction by analysing transcriptional start sites (TSS), CRISRPi identified 92% of core cell essential genes and did not show a bias to regions involved in copy number alterations. However, bidirectional promoters scored as false positives in CRISRPi. We conclude that CRISPRc and CRISPRi have different off-target effects and combining these approaches provides complementary information in loss-of-function genetic screens.

PMID:
28534478
PMCID:
PMC5457492
DOI:
10.1038/ncomms15403
[Indexed for MEDLINE]
Free PMC Article

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