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In vivo genome editing using Staphylococcus aureus Cas9

Author

Listed:
  • F. Ann Ran

    (Broad Institute of MIT and Harvard
    Society of Fellows, Harvard University)

  • Le Cong

    (Broad Institute of MIT and Harvard
    Massachusetts Institute of Technology)

  • Winston X. Yan

    (Broad Institute of MIT and Harvard
    Graduate Program in Biophysics, Harvard Medical School
    Harvard Medical School)

  • David A. Scott

    (Broad Institute of MIT and Harvard
    McGovern Institute for Brain Research, Massachusetts Institute of Technology
    Massachusetts Institute of Technology)

  • Jonathan S. Gootenberg

    (Broad Institute of MIT and Harvard
    Harvard Medical School)

  • Andrea J. Kriz

    (Massachusetts Institute of Technology)

  • Bernd Zetsche

    (Broad Institute of MIT and Harvard)

  • Ophir Shalem

    (Broad Institute of MIT and Harvard)

  • Xuebing Wu

    (David H. Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology
    Computational and Systems Biology Graduate Program, Massachusetts Institute of Technology)

  • Kira S. Makarova

    (National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health)

  • Eugene V. Koonin

    (National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health)

  • Phillip A. Sharp

    (Massachusetts Institute of Technology
    David H. Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology)

  • Feng Zhang

    (Broad Institute of MIT and Harvard
    McGovern Institute for Brain Research, Massachusetts Institute of Technology
    Massachusetts Institute of Technology
    Massachusetts Institute of Technology)

Abstract

The RNA-guided endonuclease Cas9 has emerged as a versatile genome-editing platform. However, the size of the commonly used Cas9 from Streptococcus pyogenes (SpCas9) limits its utility for basic research and therapeutic applications that use the highly versatile adeno-associated virus (AAV) delivery vehicle. Here, we characterize six smaller Cas9 orthologues and show that Cas9 from Staphylococcus aureus (SaCas9) can edit the genome with efficiencies similar to those of SpCas9, while being more than 1 kilobase shorter. We packaged SaCas9 and its single guide RNA expression cassette into a single AAV vector and targeted the cholesterol regulatory gene Pcsk9 in the mouse liver. Within one week of injection, we observed >40% gene modification, accompanied by significant reductions in serum Pcsk9 and total cholesterol levels. We further assess the genome-wide targeting specificity of SaCas9 and SpCas9 using BLESS, and demonstrate that SaCas9-mediated in vivo genome editing has the potential to be efficient and specific.

Suggested Citation

  • F. Ann Ran & Le Cong & Winston X. Yan & David A. Scott & Jonathan S. Gootenberg & Andrea J. Kriz & Bernd Zetsche & Ophir Shalem & Xuebing Wu & Kira S. Makarova & Eugene V. Koonin & Phillip A. Sharp & , 2015. "In vivo genome editing using Staphylococcus aureus Cas9," Nature, Nature, vol. 520(7546), pages 186-191, April.
    • Handle: RePEc:nat:nature:v:520:y:2015:i:7546:d:10.1038_nature14299
    DOI: 10.1038/nature14299
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