Predicting the mutations generated by repair of Cas9-induced double-strand breaks.
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Authors
Allen, Felicity
Crepaldi, Luca
Alsinet, Clara
Strong, Alexander J
De Angeli, Pietro
Páleníková, Petra
Khodak, Anton
Kosicki, Michael
Muñoz-Martínez, Francisco
Metzakopian, Emmanouil
Parts, Leopold
Publication Date
2018-11-27Journal Title
Nat Biotechnol
ISSN
1087-0156
Publisher
Springer Science and Business Media LLC
Language
eng
Type
Article
This Version
AM
Physical Medium
Print-Electronic
Metadata
Show full item recordCitation
Allen, F., Crepaldi, L., Alsinet, C., Strong, A. J., Kleshchevnikov, V., De Angeli, P., Páleníková, P., et al. (2018). Predicting the mutations generated by repair of Cas9-induced double-strand breaks.. Nat Biotechnol https://doi.org/10.1038/nbt.4317
Abstract
The DNA mutation produced by cellular repair of a CRISPR-Cas9-generated double-strand break determines its phenotypic effect. It is known that the mutational outcomes are not random, but depend on DNA sequence at the targeted location. Here we systematically study the influence of flanking DNA sequence on repair outcome by measuring the edits generated by >40,000 guide RNAs (gRNAs) in synthetic constructs. We performed the experiments in a range of genetic backgrounds and using alternative CRISPR-Cas9 reagents. In total, we gathered data for >109 mutational outcomes. The majority of reproducible mutations are insertions of a single base, short deletions or longer microhomology-mediated deletions. Each gRNA has an individual cell-line-dependent bias toward particular outcomes. We uncover sequence determinants of the mutations produced and use these to derive a predictor of Cas9 editing outcomes. Improved understanding of sequence repair will allow better design of gene editing experiments.
Sponsorship
Cancer Research UK (18796)
Wellcome Trust (206388/Z/17/Z)
Wellcome Trust (084812/Z/08/Z)
Wellcome Trust (200848/Z/16/Z)
Wellcome Trust (100140/Z/12/Z)
Identifiers
External DOI: https://doi.org/10.1038/nbt.4317
This record's URL: https://www.repository.cam.ac.uk/handle/1810/286946
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