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DNA Repair Profiling Reveals Nonrandom Outcomes at Cas9-Mediated Breaks

van Overbeek, Megan and Capurso, Daniel and Carter, Mathew M. and Thompson, Mathew S. and Frias, Elizabeth and Reece-Hoyes, John and Nye, Christopher and Gradia, Scott and Vidal, Bastien and Zheng, Jiashun and Hoffman, Gregory and Fuller, Christopher K. and May, Andrew P. (2016) DNA Repair Profiling Reveals Nonrandom Outcomes at Cas9-Mediated Breaks. Molecular Cell, 63 (4). pp. 633-646. ISSN 1097-4164

Abstract

The repair outcomes at site-specific DNA double-strand breaks (DSBs) generated by the RNA-guided DNA endonuclease Cas9 determine how gene function is altered. Despite the widespread adoption of CRISPR-Cas9 technology to induce DSBs for genome engineering, the resulting repair products have not been examined in depth. Here, the DNA repair profiles of 223 sites in the human genome demonstrate that the pattern of DNA repair following Cas9 cutting at each site is nonrandom and consistent across experimental replicates, cell lines, and reagent delivery methods. Furthermore, the repair outcomes are determined by the protospacer sequence rather than genomic context, indicating that DNA repair profiling in cell lines can be used to anticipate repair outcomes in primary cells. Chemical inhibition of DNA-PK enabled dissection of the DNA repair profiles into contributions from c-NHEJ and MMEJ. Finally, this work elucidates a strategy for using “error-prone” DNA-repair machinery to generate precise edits.

Item Type: Article
Date Deposited: 14 Mar 2018 00:45
Last Modified: 25 Jan 2019 00:45
URI: https://oak.novartis.com/id/eprint/29796

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