Increasing the efficiency of precise genome editing with CRISPR-Cas9 by inhibition of nonhomologous end joining
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Ploegh_Increasing the efficiency.pdf
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Author(s)
Maruyama, Takeshi
Dougan, Stephanie K
Truttmann, Matthias C
Bilate, Angelina M
Ingram, Jessica R
Ploegh, Hidde
Date Issued
May 2015
Journal
Nature Biotechnology
Publisher
Nature Publishing Group
Citation
Maruyama, Takeshi et al. “Increasing the Efficiency of Precise Genome Editing with CRISPR-Cas9 by Inhibition of Nonhomologous End Joining.” Nature Biotechnology 33.5 (2015): 538–542.
Version
Author's final manuscript
Abstract
Methods to introduce targeted double-strand breaks (DSBs) into DNA enable precise genome editing by increasing the rate at which externally supplied DNA fragments are incorporated into the genome through homologous recombination. The efficiency of these methods is limited by nonhomologous end joining (NHEJ), an alternative DNA repair pathway that competes with homology-directed repair (HDR). To promote HDR at the expense of NHEJ, we targeted DNA ligase IV, a key enzyme in the NHEJ pathway, using the inhibitor Scr7. Scr7 treatment increased the efficiency of HDR-mediated genome editing, using Cas9 in mammalian cell lines and in mice for all four genes examined, up to 19-fold. This approach should be applicable to other customizable endonucleases, such as zinc finger nucleases and transcription activator–like effector nucleases, and to nonmammalian cells with sufficiently conserved mechanisms of NHEJ and HDR.
MIT Department
Massachusetts Institute of Technology. Department of Biology
Whitehead Institute for Biomedical Research
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Creative Commons Attribution-Noncommercial-Share Alike
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DOI of Published Version
https://doi.org/10.1038/nbt.3190
