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Whole genomic analysis reveals atypical non-homologous off-target large structural variants induced by CRISPR-Cas9-mediated genome editing

Author

Listed:
  • Hsiu-Hui Tsai

    (Chang Gung Memorial Hospital at Linkou)

  • Hsiao-Jung Kao

    (Academia Sinica)

  • Ming-Wei Kuo

    (Chang Gung Memorial Hospital at Linkou)

  • Chin-Hsien Lin

    (National Taiwan University Hospital and School of Medicine)

  • Chun-Min Chang

    (Chang Gung Memorial Hospital at Linkou)

  • Yi-Yin Chen

    (Chang Gung Memorial Hospital at Linkou)

  • Hsiao-Huei Chen

    (Academia Sinica)

  • Pui-Yan Kwok

    (Academia Sinica
    University of California)

  • Alice L. Yu

    (Chang Gung Memorial Hospital at Linkou
    University of California
    Academia Sinica)

  • John Yu

    (Chang Gung Memorial Hospital at Linkou
    Academia Sinica)

Abstract

CRISPR-Cas9 genome editing has promising therapeutic potential for genetic diseases and cancers, but safety could be a concern. Here we use whole genomic analysis by 10x linked-read sequencing and optical genome mapping to interrogate the genome integrity after editing and in comparison to four parental cell lines. In addition to the previously reported large structural variants at on-target sites, we identify heretofore unexpected large chromosomal deletions (91.2 and 136 Kb) at atypical non-homologous off-target sites without sequence similarity to the sgRNA in two edited lines. The observed large structural variants induced by CRISPR-Cas9 editing in dividing cells may result in pathogenic consequences and thus limit the usefulness of the CRISPR-Cas9 editing system for disease modeling and gene therapy. In this work, our whole genomic analysis may provide a valuable strategy to ensure genome integrity after genomic editing to minimize the risk of unintended effects in research and clinical applications.

Suggested Citation

  • Hsiu-Hui Tsai & Hsiao-Jung Kao & Ming-Wei Kuo & Chin-Hsien Lin & Chun-Min Chang & Yi-Yin Chen & Hsiao-Huei Chen & Pui-Yan Kwok & Alice L. Yu & John Yu, 2023. "Whole genomic analysis reveals atypical non-homologous off-target large structural variants induced by CRISPR-Cas9-mediated genome editing," Nature Communications, Nature, vol. 14(1), pages 1-9, December.
    • Handle: RePEc:nat:natcom:v:14:y:2023:i:1:d:10.1038_s41467-023-40901-x
    DOI: 10.1038/s41467-023-40901-x
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    1. Ida Höijer & Anastasia Emmanouilidou & Rebecka Östlund & Robin Schendel & Selma Bozorgpana & Marcel Tijsterman & Lars Feuk & Ulf Gyllensten & Marcel Hoed & Adam Ameur, 2022. "CRISPR-Cas9 induces large structural variants at on-target and off-target sites in vivo that segregate across generations," Nature Communications, Nature, vol. 13(1), pages 1-10, December.
    2. Stamatis Papathanasiou & Styliani Markoulaki & Logan J. Blaine & Mitchell L. Leibowitz & Cheng-Zhong Zhang & Rudolf Jaenisch & David Pellman, 2021. "Whole chromosome loss and genomic instability in mouse embryos after CRISPR-Cas9 genome editing," Nature Communications, Nature, vol. 12(1), pages 1-7, December.
    3. Shiran Abadi & Winston X Yan & David Amar & Itay Mayrose, 2017. "A machine learning approach for predicting CRISPR-Cas9 cleavage efficiencies and patterns underlying its mechanism of action," PLOS Computational Biology, Public Library of Science, vol. 13(10), pages 1-24, October.
    4. Grégoire Cullot & Julian Boutin & Jérôme Toutain & Florence Prat & Perrine Pennamen & Caroline Rooryck & Martin Teichmann & Emilie Rousseau & Isabelle Lamrissi-Garcia & Véronique Guyonnet-Duperat & Al, 2019. "CRISPR-Cas9 genome editing induces megabase-scale chromosomal truncations," Nature Communications, Nature, vol. 10(1), pages 1-14, December.
    5. M. Kyle Cromer & Valentin V. Barsan & Erich Jaeger & Mengchi Wang & Jessica P. Hampton & Feng Chen & Drew Kennedy & Jenny Xiao & Irina Khrebtukova & Ana Granat & Tiffany Truong & Matthew H. Porteus, 2022. "Ultra-deep sequencing validates safety of CRISPR/Cas9 genome editing in human hematopoietic stem and progenitor cells," Nature Communications, Nature, vol. 13(1), pages 1-11, December.
    6. Lucas Pelkmans & Eugenio Fava & Hannes Grabner & Michael Hannus & Bianca Habermann & Eberhard Krausz & Marino Zerial, 2005. "Genome-wide analysis of human kinases in clathrin- and caveolae/raft-mediated endocytosis," Nature, Nature, vol. 436(7047), pages 78-86, July.
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