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Comparative optimization of combinatorial CRISPR screens

Author

Listed:
  • Ruitong Li

    (Broad Institute of Harvard and MIT)

  • Olaf Klingbeil

    (Cold Spring Harbor Laboratory)

  • Davide Monducci

    (Broad Institute of Harvard and MIT)

  • Michael J. Young

    (Broad Institute of Harvard and MIT)

  • Diego J. Rodriguez

    (Broad Institute of Harvard and MIT)

  • Zaid Bayyat

    (Broad Institute of Harvard and MIT)

  • Joshua M. Dempster

    (Broad Institute of Harvard and MIT)

  • Devishi Kesar

    (Broad Institute of Harvard and MIT)

  • Xiaoping Yang

    (Broad Institute of Harvard and MIT)

  • Mahdi Zamanighomi

    (Broad Institute of Harvard and MIT)

  • Christopher R. Vakoc

    (Cold Spring Harbor Laboratory)

  • Takahiro Ito

    (Broad Institute of Harvard and MIT
    Harvard Medical School
    Dana-Farber Cancer Institute
    Scorpion Therapeutics)

  • William R. Sellers

    (Broad Institute of Harvard and MIT
    Harvard Medical School
    Dana-Farber Cancer Institute)

Abstract

Combinatorial CRISPR technologies have emerged as a transformative approach to systematically probe genetic interactions and dependencies of redundant gene pairs. However, the performance of different functional genomic tools for multiplexing sgRNAs vary widely. Here, we generate and benchmark ten distinct pooled combinatorial CRISPR libraries targeting paralog pairs to optimize digenic knockout screens. Libraries composed of dual Streptococcus pyogenes Cas9 (spCas9), orthogonal spCas9 and Staphylococcus aureus (saCas9), and enhanced Cas12a from Acidaminococcus were evaluated. We demonstrate a combination of alternative tracrRNA sequences from spCas9 consistently show superior effect size and positional balance between the sgRNAs as a robust combinatorial approach to profile genetic interactions of multiple genes.

Suggested Citation

  • Ruitong Li & Olaf Klingbeil & Davide Monducci & Michael J. Young & Diego J. Rodriguez & Zaid Bayyat & Joshua M. Dempster & Devishi Kesar & Xiaoping Yang & Mahdi Zamanighomi & Christopher R. Vakoc & Ta, 2022. "Comparative optimization of combinatorial CRISPR screens," Nature Communications, Nature, vol. 13(1), pages 1-10, December.
    • Handle: RePEc:nat:natcom:v:13:y:2022:i:1:d:10.1038_s41467-022-30196-9
    DOI: 10.1038/s41467-022-30196-9
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