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Efficient single-copy HDR by 5' modified long dsDNA donors

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Cite this article as: eLife 2018;7:e39468 doi: 10.7554/eLife.39468

Abstract

CRISPR/Cas9 efficiently induces targeted mutations via non-homologous-end-joining but for genome editing, precise, homology-directed repair (HDR) of endogenous DNA stretches is a prerequisite. To favor HDR, many approaches interfere with the repair machinery or manipulate Cas9 itself. Using Medaka we show that the modification of 5' ends of long dsDNA donors strongly enhances HDR, favors efficient single-copy integration by retaining a monomeric donor conformation thus facilitating successful gene replacement or tagging.

Article and author information

Author details

  1. Jose Arturo Gutierrez-Triana

    Centre for Organismal Studies, Heidelberg University, Heidelberg, Germany
    Competing interests
    The authors declare that no competing interests exist.
  2. Tinatini Tavhelidse

    Centre for Organismal Studies, Heidelberg University, Heidelberg, Germany
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-6103-9019
  3. Thomas Thumberger

    Centre for Organismal Studies, Heidelberg University, Heidelberg, Germany
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-8485-457X
  4. Isabelle Thomas

    Centre for Organismal Studies, Heidelberg University, Heidelberg, Germany
    Competing interests
    The authors declare that no competing interests exist.
  5. Beate Wittbrodt

    Centre for Organismal Studies, Heidelberg University, Heidelberg, Germany
    Competing interests
    The authors declare that no competing interests exist.
  6. Tanja Kellner

    Centre for Organismal Studies, Heidelberg University, Heidelberg, Germany
    Competing interests
    The authors declare that no competing interests exist.
  7. Kerim Anlas

    Centre for Organismal Studies, Heidelberg University, Heidelberg, Germany
    Competing interests
    The authors declare that no competing interests exist.
  8. Erika Tsingos

    Centre for Organismal Studies, Heidelberg University, Heidelberg, Germany
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-7267-160X
  9. Joachim Wittbrodt

    Centre for Organismal Studies, Heidelberg University, Heidelberg, Germany
    For correspondence
    jochen.wittbrodt@cos.uni-heidelberg.de
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-8550-7377

Funding

Deutsche Forschungsgemeinschaft (CRC 873 project A3)

  • Joachim Wittbrodt

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Reviewing Editor

  1. Alejandro Sánchez Alvarado, Stowers Institute for Medical Research, United States

Publication history

  1. Received: June 22, 2018
  2. Accepted: August 14, 2018
  3. Accepted Manuscript published: August 29, 2018 (version 1)
  4. Version of Record published: September 5, 2018 (version 2)

Copyright

© 2018, Gutierrez-Triana et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

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