Skip to main content

Compact and highly active next-generation libraries for CRISPR-mediated gene repression and activation.

Author(s): Horlbeck, Max A; Gilbert, Luke A; Villalta, Jacqueline E; Adamson, Britt; Pak, Ryan A; et al

To refer to this page use:
Abstract: We recently found that nucleosomes directly block access of CRISPR/Cas9 to DNA (Horlbeck et al., 2016). Here, we build on this observation with a comprehensive algorithm that incorporates chromatin, position, and sequence features to accurately predict highly effective single guide RNAs (sgRNAs) for targeting nuclease-dead Cas9-mediated transcriptional repression (CRISPRi) and activation (CRISPRa). We use this algorithm to design next-generation genome-scale CRISPRi and CRISPRa libraries targeting human and mouse genomes. A CRISPRi screen for essential genes in K562 cells demonstrates that the large majority of sgRNAs are highly active. We also find CRISPRi does not exhibit any detectable non-specific toxicity recently observed with CRISPR nuclease approaches. Precision-recall analysis shows that we detect over 90% of essential genes with minimal false positives using a compact 5 sgRNA/gene library. Our results establish CRISPRi and CRISPRa as premier tools for loss- or gain-of-function studies and provide a general strategy for identifying Cas9 target sites.
Publication Date: 23-Sep-2016
Citation: Horlbeck, Max A, Gilbert, Luke A, Villalta, Jacqueline E, Adamson, Britt, Pak, Ryan A, Chen, Yuwen, Fields, Alexander P, Park, Chong Yon, Corn, Jacob E, Kampmann, Martin, Weissman, Jonathan S. (2016). Compact and highly active next-generation libraries for CRISPR-mediated gene repression and activation.. eLife, 5 (10.7554/eLife.19760
DOI: doi:10.7554/eLife.19760
ISSN: 2050-084X
EISSN: 2050-084X
Pages: 1 - 20
Language: eng
Type of Material: Journal Article
Journal/Proceeding Title: eLife
Version: Final published version. This is an open access article.

Items in OAR@Princeton are protected by copyright, with all rights reserved, unless otherwise indicated.