Optimized libraries for CRISPR-Cas9 genetic screens with multiple modalities

Kendall R. Sanson, Ruth E. Hanna, Mudra Hegde, Katherine F. Donovan, Christine Strand, Meagan E. Sullender, Emma W. Vaimberg, Amy Goodale, David E. Root, Federica Piccioni and John G. Doench ()
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Kendall R. Sanson: Broad Institute of Harvard and MIT
Ruth E. Hanna: Broad Institute of Harvard and MIT
Mudra Hegde: Broad Institute of Harvard and MIT
Katherine F. Donovan: Broad Institute of Harvard and MIT
Christine Strand: Broad Institute of Harvard and MIT
Meagan E. Sullender: Broad Institute of Harvard and MIT
Emma W. Vaimberg: Broad Institute of Harvard and MIT
Amy Goodale: Broad Institute of Harvard and MIT
David E. Root: Broad Institute of Harvard and MIT
Federica Piccioni: Broad Institute of Harvard and MIT
John G. Doench: Broad Institute of Harvard and MIT

Nature Communications, 2018, vol. 9, issue 1, 1-15

Abstract: Abstract The creation of genome-wide libraries for CRISPR knockout (CRISPRko), interference (CRISPRi), and activation (CRISPRa) has enabled the systematic interrogation of gene function. Here, we show that our recently-described CRISPRko library (Brunello) is more effective than previously published libraries at distinguishing essential and non-essential genes, providing approximately the same perturbation-level performance improvement over GeCKO libraries as GeCKO provided over RNAi. Additionally, we present genome-wide libraries for CRISPRi (Dolcetto) and CRISPRa (Calabrese), and show in negative selection screens that Dolcetto, with fewer sgRNAs per gene, outperforms existing CRISPRi libraries and achieves comparable performance to CRISPRko in detecting essential genes. We also perform positive selection CRISPRa screens and demonstrate that Calabrese outperforms the SAM approach at identifying vemurafenib resistance genes. We further compare CRISPRa to genome-scale libraries of open reading frames (ORFs). Together, these libraries represent a suite of genome-wide tools to efficiently interrogate gene function with multiple modalities.

Date: 2018
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DOI: 10.1038/s41467-018-07901-8

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