Complementary information derived from CRISPR Cas9 mediated gene deletion and suppression

Joseph Rosenbluh, Han Xu, William Harrington, Stanley Gill, Xiaoxing Wang, Francisca Vazquez, David E. Root, Aviad Tsherniak, William Hahn

Research output: Contribution to journalArticleResearchpeer-review

26 Citations (Scopus)

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.

Original languageEnglish
Article number15403
Number of pages8
JournalNature Communications
Volume8
DOIs
Publication statusPublished - 23 May 2017

Keywords

  • cancer genomics
  • CRISPR-Cas9 genome editing
  • functional genomics

Cite this

Rosenbluh, Joseph ; Xu, Han ; Harrington, William ; Gill, Stanley ; Wang, Xiaoxing ; Vazquez, Francisca ; Root, David E. ; Tsherniak, Aviad ; Hahn, William. / Complementary information derived from CRISPR Cas9 mediated gene deletion and suppression. In: Nature Communications. 2017 ; Vol. 8.
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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.",
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Rosenbluh, J, Xu, H, Harrington, W, Gill, S, Wang, X, Vazquez, F, Root, DE, Tsherniak, A & Hahn, W 2017, 'Complementary information derived from CRISPR Cas9 mediated gene deletion and suppression', Nature Communications, vol. 8, 15403. https://doi.org/10.1038/ncomms15403

Complementary information derived from CRISPR Cas9 mediated gene deletion and suppression. / Rosenbluh, Joseph; Xu, Han; Harrington, William; Gill, Stanley; Wang, Xiaoxing; Vazquez, Francisca; Root, David E.; Tsherniak, Aviad; Hahn, William.

In: Nature Communications, Vol. 8, 15403, 23.05.2017.

Research output: Contribution to journalArticleResearchpeer-review

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AU - Root, David E.

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