IDEAS home Printed from https://ideas.repec.org/a/nat/natcom/v9y2018i1d10.1038_s41467-018-05477-x.html
   My bibliography  Save this article

Directed evolution of CRISPR-Cas9 to increase its specificity

Author

Listed:
  • Jungjoon K. Lee

    (Toolgen)

  • Euihwan Jeong

    (Institute for Basic Science (IBS)
    Seoul National University)

  • Joonsun Lee

    (Toolgen)

  • Minhee Jung

    (Toolgen)

  • Eunji Shin

    (Toolgen)

  • Young-hoon Kim

    (Toolgen)

  • Kangin Lee

    (Toolgen)

  • Inyoung Jung

    (Toolgen)

  • Daesik Kim

    (Seoul National University)

  • Seokjoong Kim

    (Toolgen)

  • Jin-Soo Kim

    (Institute for Basic Science (IBS)
    Seoul National University)

Abstract

The use of CRISPR-Cas9 as a therapeutic reagent is hampered by its off-target effects. Although rationally designed S. pyogenes Cas9 (SpCas9) variants that display higher specificities than the wild-type SpCas9 protein are available, these attenuated Cas9 variants are often poorly efficient in human cells. Here, we develop a directed evolution approach in E. coli to obtain Sniper-Cas9, which shows high specificities without killing on-target activities in human cells. Unlike other engineered Cas9 variants, Sniper-Cas9 shows WT-level on-target activities with extended or truncated sgRNAs with further reduced off-target activities and works well in a preassembled ribonucleoprotein (RNP) format to allow DNA-free genome editing.

Suggested Citation

  • Jungjoon K. Lee & Euihwan Jeong & Joonsun Lee & Minhee Jung & Eunji Shin & Young-hoon Kim & Kangin Lee & Inyoung Jung & Daesik Kim & Seokjoong Kim & Jin-Soo Kim, 2018. "Directed evolution of CRISPR-Cas9 to increase its specificity," Nature Communications, Nature, vol. 9(1), pages 1-10, December.
    • Handle: RePEc:nat:natcom:v:9:y:2018:i:1:d:10.1038_s41467-018-05477-x
    DOI: 10.1038/s41467-018-05477-x
    as

    Download full text from publisher

    File URL: https://www.nature.com/articles/s41467-018-05477-x
    File Function: Abstract
    Download Restriction: no
    File URL: https://libkey.io/10.1038/s41467-018-05477-x?utm_source=ideas
    LibKey link: if access is restricted and if your library uses this service, LibKey will redirect you to where you can use your library subscription to access this item
    ---><---

    Citations

    Citations are extracted by the CitEc Project, subscribe to its RSS feed for this item.
    as


    Cited by:

    1. Dawn G. L. Thean & Hoi Yee Chu & John H. C. Fong & Becky K. C. Chan & Peng Zhou & Cynthia C. S. Kwok & Yee Man Chan & Silvia Y. L. Mak & Gigi C. G. Choi & Joshua W. K. Ho & Zongli Zheng & Alan S. L. W, 2022. "Machine learning-coupled combinatorial mutagenesis enables resource-efficient engineering of CRISPR-Cas9 genome editor activities," Nature Communications, Nature, vol. 13(1), pages 1-14, December.
    2. Péter István Kulcsár & András Tálas & Zoltán Ligeti & Sarah Laura Krausz & Ervin Welker, 2022. "SuperFi-Cas9 exhibits remarkable fidelity but severely reduced activity yet works effectively with ABE8e," Nature Communications, Nature, vol. 13(1), pages 1-11, December.
    3. Jianli Tao & Daniel E. Bauer & Roberto Chiarle, 2023. "Assessing and advancing the safety of CRISPR-Cas tools: from DNA to RNA editing," Nature Communications, Nature, vol. 14(1), pages 1-16, December.
    4. Péter István Kulcsár & András Tálas & Zoltán Ligeti & Eszter Tóth & Zsófia Rakvács & Zsuzsa Bartos & Sarah Laura Krausz & Ágnes Welker & Vanessza Laura Végi & Krisztina Huszár & Ervin Welker, 2023. "A cleavage rule for selection of increased-fidelity SpCas9 variants with high efficiency and no detectable off-targets," Nature Communications, Nature, vol. 14(1), pages 1-20, December.
    5. Burcu Bestas & Sandra Wimberger & Dmitrii Degtev & Alexandra Madsen & Antje K. Rottner & Fredrik Karlsson & Sergey Naumenko & Megan Callahan & Julia Liz Touza & Margherita Francescatto & Carl Ivar Möl, 2023. "A Type II-B Cas9 nuclease with minimized off-targets and reduced chromosomal translocations in vivo," Nature Communications, Nature, vol. 14(1), pages 1-15, December.
    6. Pierre Aldag & Marius Rutkauskas & Julene Madariaga-Marcos & Inga Songailiene & Tomas Sinkunas & Felix Kemmerich & Dominik Kauert & Virginijus Siksnys & Ralf Seidel, 2023. "Dynamic interplay between target search and recognition for a Type I CRISPR-Cas system," Nature Communications, Nature, vol. 14(1), pages 1-14, December.
    7. Yanbo Wang & W. Taylor Cottle & Haobo Wang & Momcilo Gavrilov & Roger S. Zou & Minh-Tam Pham & Srinivasan Yegnasubramanian & Scott Bailey & Taekjip Ha, 2022. "Achieving single nucleotide sensitivity in direct hybridization genome imaging," Nature Communications, Nature, vol. 13(1), pages 1-12, December.
    8. András Tálas & Dorottya A. Simon & Péter I. Kulcsár & Éva Varga & Sarah L. Krausz & Ervin Welker, 2021. "BEAR reveals that increased fidelity variants can successfully reduce the mismatch tolerance of adenine but not cytosine base editors," Nature Communications, Nature, vol. 12(1), pages 1-14, December.
    9. Akiko Tomita & Hiroyuki Sasanuma & Tomoo Owa & Yuka Nakazawa & Mayuko Shimada & Takahiro Fukuoka & Tomoo Ogi & Shinichiro Nakada, 2023. "Inducing multiple nicks promotes interhomolog homologous recombination to correct heterozygous mutations in somatic cells," Nature Communications, Nature, vol. 14(1), pages 1-20, December.
    10. Giulia I. Corsi & Kunli Qu & Ferhat Alkan & Xiaoguang Pan & Yonglun Luo & Jan Gorodkin, 2022. "CRISPR/Cas9 gRNA activity depends on free energy changes and on the target PAM context," Nature Communications, Nature, vol. 13(1), pages 1-14, December.

    More about this item

    Statistics

    Access and download statistics

    Corrections

    All material on this site has been provided by the respective publishers and authors. You can help correct errors and omissions. When requesting a correction, please mention this item's handle: RePEc:nat:natcom:v:9:y:2018:i:1:d:10.1038_s41467-018-05477-x. See general information about how to correct material in RePEc.

    If you have authored this item and are not yet registered with RePEc, we encourage you to do it here. This allows to link your profile to this item. It also allows you to accept potential citations to this item that we are uncertain about.

    We have no bibliographic references for this item. You can help adding them by using this form .

    If you know of missing items citing this one, you can help us creating those links by adding the relevant references in the same way as above, for each refering item. If you are a registered author of this item, you may also want to check the "citations" tab in your RePEc Author Service profile, as there may be some citations waiting for confirmation.

    For technical questions regarding this item, or to correct its authors, title, abstract, bibliographic or download information, contact: Sonal Shukla or Springer Nature Abstracting and Indexing (email available below). General contact details of provider: http://www.nature.com .

    Please note that corrections may take a couple of weeks to filter through the various RePEc services.

    IDEAS is a RePEc service. RePEc uses bibliographic data supplied by the respective publishers.