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Mechanistic and genetic basis of single-strand templated repair at Cas12a-induced DNA breaks in Chlamydomonas reinhardtii

Author

Listed:
  • Aron Ferenczi

    (University of Edinburgh)

  • Yen Peng Chew

    (University of Edinburgh)

  • Erika Kroll

    (University of Edinburgh
    Rothamsted Research)

  • Charlotte Koppenfels

    (University of Edinburgh)

  • Andrew Hudson

    (University of Edinburgh)

  • Attila Molnar

    (University of Edinburgh)

Abstract

Single-stranded oligodeoxynucleotides (ssODNs) are widely used as DNA repair templates in CRISPR/Cas precision genome editing. However, the underlying mechanisms of single-strand templated DNA repair (SSTR) are inadequately understood, constraining rational improvements to precision editing. Here we study SSTR at CRISPR/Cas12a-induced DNA double-strand breaks (DSBs) in the eukaryotic model green microalga Chlamydomonas reinhardtii. We demonstrate that ssODNs physically incorporate into the genome during SSTR at Cas12a-induced DSBs. This process is genetically independent of the Rad51-dependent homologous recombination and Fanconi anemia pathways, is strongly antagonized by non-homologous end-joining, and is mediated almost entirely by the alternative end-joining enzyme polymerase θ. These findings suggest differences in SSTR between C. reinhardtii and animals. Our work illustrates the promising potentially of C. reinhardtii as a model organism for studying nuclear DNA repair.

Suggested Citation

  • Aron Ferenczi & Yen Peng Chew & Erika Kroll & Charlotte Koppenfels & Andrew Hudson & Attila Molnar, 2021. "Mechanistic and genetic basis of single-strand templated repair at Cas12a-induced DNA breaks in Chlamydomonas reinhardtii," Nature Communications, Nature, vol. 12(1), pages 1-12, December.
    • Handle: RePEc:nat:natcom:v:12:y:2021:i:1:d:10.1038_s41467-021-27004-1
    DOI: 10.1038/s41467-021-27004-1
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    References listed on IDEAS

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