Pharmacological inhibition of DNA-PK stimulates Cas9-mediated genome editing View Full Text


Ontology type: schema:ScholarlyArticle      Open Access: True


Article Info

DATE

2015-08-27

AUTHORS

Francis Robert, Mathilde Barbeau, Sylvain Éthier, Josée Dostie, Jerry Pelletier

ABSTRACT

BackgroundThe ability to modify the genome of any cell at a precise location has drastically improved with the recent discovery and implementation of CRISPR/Cas9 editing technology. However, the capacity to introduce specific directed changes at given loci is hampered by the fact that the major cellular repair pathway that occurs following Cas9-mediated DNA cleavage is the erroneous non-homologous end joining (NHEJ) pathway. Homology-directed recombination (HDR) is far less efficient than NHEJ and makes screening of clones containing directed changes time-consuming and labor-intensive.MethodsWe investigated the possibility of pharmacologically inhibiting DNA-PKcs, a key player in NHEJ, using small molecule inhibitors (NU7441 and KU-0060648), to ameliorate the rates of HDR repair events. These compounds were tested in a sensitive reporter assay capable of simultaneously informing on NHEJ and HDR, as well as on an endogenous gene targeted by Cas9.ResultsWe find that NU7441 and KU-0060648 reduce the frequency of NHEJ while increasing the rate of HDR following Cas9-mediated DNA cleavage.ConclusionsOur results identify two small molecules compatible for use with Cas9-editing technology to improve the frequency of HDR. More... »

PAGES

93

Identifiers

URI

http://scigraph.springernature.com/pub.10.1186/s13073-015-0215-6

DOI

http://dx.doi.org/10.1186/s13073-015-0215-6

DIMENSIONS

https://app.dimensions.ai/details/publication/pub.1031597914

PUBMED

https://www.ncbi.nlm.nih.gov/pubmed/26307031


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