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CRISPR-Cas12a induced DNA double-strand breaks are repaired by multiple pathways with different mutation profiles in Magnaporthe oryzae

Author

Listed:
  • Jun Huang

    (Kansas State University)

  • David Rowe

    (Kansas State University)

  • Pratima Subedi

    (Kansas State University)

  • Wei Zhang

    (Kansas State University)

  • Tyler Suelter

    (Kansas State University)

  • Barbara Valent

    (Kansas State University)

  • David E. Cook

    (Kansas State University)

Abstract

CRISPR-Cas mediated genome engineering has revolutionized functional genomics. However, understanding of DNA repair following Cas-mediated DNA cleavage remains incomplete. Using Cas12a ribonucleoprotein genome editing in the fungal pathogen, Magnaporthe oryzae, we detail non-canonical DNA repair outcomes from hundreds of transformants. Sanger and nanopore sequencing analysis reveals significant variation in DNA repair profiles, ranging from small INDELs to kilobase size deletions and insertions. Furthermore, we find the frequency of DNA repair outcomes varies between loci. The results are not specific to the Cas-nuclease or selection procedure. Through Ku80 deletion analysis, a key protein required for canonical non-homologous end joining, we demonstrate activity of an alternative end joining mechanism that creates larger DNA deletions, and uses longer microhomology compared to C-NHEJ. Together, our results suggest preferential DNA repair pathway activity in the genome that can create different mutation profiles following repair, which could create biased genome variation and impact genome engineering and genome evolution.

Suggested Citation

  • Jun Huang & David Rowe & Pratima Subedi & Wei Zhang & Tyler Suelter & Barbara Valent & David E. Cook, 2022. "CRISPR-Cas12a induced DNA double-strand breaks are repaired by multiple pathways with different mutation profiles in Magnaporthe oryzae," Nature Communications, Nature, vol. 13(1), pages 1-18, December.
    • Handle: RePEc:nat:natcom:v:13:y:2022:i:1:d:10.1038_s41467-022-34736-1
    DOI: 10.1038/s41467-022-34736-1
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    1. Fatwa Adikusuma & Sandra Piltz & Mark A. Corbett & Michelle Turvey & Shaun R. McColl & Karla J. Helbig & Michael R. Beard & James Hughes & Richard T. Pomerantz & Paul Q. Thomas, 2018. "Large deletions induced by Cas9 cleavage," Nature, Nature, vol. 560(7717), pages 8-9, August.
    2. James E. Haber, 1999. "Gatekeepers of recombination," Nature, Nature, vol. 398(6729), pages 665-667, April.
    3. Robin van Schendel & Sophie F. Roerink & Vincent Portegijs & Sander van den Heuvel & Marcel Tijsterman, 2015. "Polymerase Θ is a key driver of genome evolution and of CRISPR/Cas9-mediated mutagenesis," Nature Communications, Nature, vol. 6(1), pages 1-8, November.
    4. Keiichiro Suzuki & Yuji Tsunekawa & Reyna Hernandez-Benitez & Jun Wu & Jie Zhu & Euiseok J. Kim & Fumiyuki Hatanaka & Mako Yamamoto & Toshikazu Araoka & Zhe Li & Masakazu Kurita & Tomoaki Hishida & Mo, 2016. "In vivo genome editing via CRISPR/Cas9 mediated homology-independent targeted integration," Nature, Nature, vol. 540(7631), pages 144-149, December.
    5. Santiago Gisler & Joana P. Gonçalves & Waseem Akhtar & Johann de Jong & Alexey V. Pindyurin & Lodewyk F. A. Wessels & Maarten van Lohuizen, 2019. "Multiplexed Cas9 targeting reveals genomic location effects and gRNA-based staggered breaks influencing mutation efficiency," Nature Communications, Nature, vol. 10(1), pages 1-14, December.
    6. John R. Walker & Richard A. Corpina & Jonathan Goldberg, 2001. "Structure of the Ku heterodimer bound to DNA and its implications for double-strand break repair," Nature, Nature, vol. 412(6847), pages 607-614, August.
    7. Ralph A. Dean & Nicholas J. Talbot & Daniel J. Ebbole & Mark L. Farman & Thomas K. Mitchell & Marc J. Orbach & Michael Thon & Resham Kulkarni & Jin-Rong Xu & Huaqin Pan & Nick D. Read & Yong-Hwan Lee , 2005. "The genome sequence of the rice blast fungus Magnaporthe grisea," Nature, Nature, vol. 434(7036), pages 980-986, April.
    8. Anne Bothmer & Tanushree Phadke & Luis A. Barrera & Carrie M Margulies & Christina S. Lee & Frank Buquicchio & Sean Moss & Hayat S. Abdulkerim & William Selleck & Hariharan Jayaram & Vic E. Myer & Cec, 2017. "Characterization of the interplay between DNA repair and CRISPR/Cas9-induced DNA lesions at an endogenous locus," Nature Communications, Nature, vol. 8(1), pages 1-12, April.
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