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Optimized libraries for CRISPR-Cas9 genetic screens with multiple modalities

Author

Listed:
  • 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)

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.

Suggested Citation

  • 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 & John G. Doench, 2018. "Optimized libraries for CRISPR-Cas9 genetic screens with multiple modalities," Nature Communications, Nature, vol. 9(1), pages 1-15, December.
    • Handle: RePEc:nat:natcom:v:9:y:2018:i:1:d:10.1038_s41467-018-07901-8
    DOI: 10.1038/s41467-018-07901-8
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    2. Peter C. DeWeirdt & Abby V. McGee & Fengyi Zheng & Ifunanya Nwolah & Mudra Hegde & John G. Doench, 2022. "Accounting for small variations in the tracrRNA sequence improves sgRNA activity predictions for CRISPR screening," Nature Communications, Nature, vol. 13(1), pages 1-11, December.
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    17. Luke Hoberecht & Pirunthan Perampalam & Aaron Lun & Jean-Philippe Fortin, 2022. "A comprehensive Bioconductor ecosystem for the design of CRISPR guide RNAs across nucleases and technologies," Nature Communications, Nature, vol. 13(1), pages 1-20, December.
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