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The interplay of DNA repair context with target sequence predictably biases Cas9-generated mutations

Author

Listed:
  • Ananth Pallaseni

    (Wellcome Genome Campus)

  • Elin Madli Peets

    (Wellcome Genome Campus)

  • Gareth Girling

    (Wellcome Genome Campus)

  • Luca Crepaldi

    (Wellcome Genome Campus)

  • Ivan Kuzmin

    (University of Tartu)

  • Marilin Moor

    (University of Tartu)

  • Núria Muñoz-Subirana

    (Leiden University Medical Center)

  • Joost Schimmel

    (Leiden University Medical Center)

  • Özdemirhan Serçin

    (BioMed X Institute (GmbH))

  • Balca R. Mardin

    (BioMed X Institute (GmbH)
    Merck Healthcare KGaA)

  • Marcel Tijsterman

    (Leiden University Medical Center
    Leiden University)

  • Hedi Peterson

    (University of Tartu)

  • Michael Kosicki

    (University of Cambridge
    Lawrence Berkeley National Laboratory)

  • Leopold Parts

    (Wellcome Genome Campus
    University of Tartu)

Abstract

Repair of double-stranded breaks generated by CRISPR/Cas9 is highly dependent on the flanking DNA sequence. To learn about interactions between DNA repair and target sequence, we measure frequencies of over 236,000 distinct Cas9-generated mutational outcomes at over 2800 synthetic target sequences in 18 DNA repair deficient mouse embryonic stem cells lines. We classify the outcomes in an unbiased way, finding a specialised role for Prkdc (DNA-PKcs protein) and Polm in creating 1 bp insertions matching the nucleotide on the protospacer-adjacent motif side of the break, a variable involvement of Nbn and Polq in the creation of different deletion outcomes, and uni-directional deletions dependent on both end-protection and end-resection. Using our dataset, we build predictive models of the mutagenic outcomes of Cas9 scission that outperform the current standards. This work improves our understanding of DNA repair gene function, and provides avenues for more precise modulation of Cas9-generated mutations.

Suggested Citation

  • Ananth Pallaseni & Elin Madli Peets & Gareth Girling & Luca Crepaldi & Ivan Kuzmin & Marilin Moor & Núria Muñoz-Subirana & Joost Schimmel & Özdemirhan Serçin & Balca R. Mardin & Marcel Tijsterman & He, 2024. "The interplay of DNA repair context with target sequence predictably biases Cas9-generated mutations," Nature Communications, Nature, vol. 15(1), pages 1-14, December.
    • Handle: RePEc:nat:natcom:v:15:y:2024:i:1:d:10.1038_s41467-024-54566-7
    DOI: 10.1038/s41467-024-54566-7
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    References listed on IDEAS

    as
    1. Michael Kosicki & Felicity Allen & Frances Steward & Kärt Tomberg & Yangyang Pan & Allan Bradley, 2022. "Cas9-induced large deletions and small indels are controlled in a convergent fashion," Nature Communications, Nature, vol. 13(1), pages 1-11, December.
    2. Joost Schimmel & Núria Muñoz-Subirana & Hanneke Kool & Robin Schendel & Marcel Tijsterman, 2021. "Small tandem DNA duplications result from CST-guided Pol α-primase action at DNA break termini," Nature Communications, Nature, vol. 12(1), pages 1-11, December.
    Full references (including those not matched with items on IDEAS)

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