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DNA mismatch repair and oligonucleotide end-protection promote base-pair substitution distal from a CRISPR/Cas9-induced DNA break.

ABSTRACT: Single-stranded oligodeoxyribonucleotide (ssODN)-mediated repair of CRISPR/Cas9-induced DNA double-strand breaks (DSB) can effectively be used to introduce small genomic alterations in a defined locus. Here, we reveal DNA mismatch repair (MMR) activity is crucial for efficient nucleotide substitution distal from the Cas9-induced DNA break when the substitution is instructed by the 3' half of the ssODN. Furthermore, protecting the ssODN 3' end with phosphorothioate linkages enhances MMR-dependent gene editing events. Our findings can be exploited to optimize efficiencies of nucleotide substitutions distal from the DSB and imply that oligonucleotide-mediated gene editing is effectuated by templated break repair.

SUBMITTER: Harmsen T

PROVIDER: S-EPMC5888797 | biostudies-other | 2018 Apr

REPOSITORIES: biostudies-other

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DNA mismatch repair and oligonucleotide end-protection promote base-pair substitution distal from a CRISPR/Cas9-induced DNA break.

Harmsen Tim T Klaasen Sjoerd S van de Vrugt Henri H Te Riele Hein H

Nucleic acids research 20180401 6

Single-stranded oligodeoxyribonucleotide (ssODN)-mediated repair of CRISPR/Cas9-induced DNA double-strand breaks (DSB) can effectively be used to introduce small genomic alterations in a defined locus. Here, we reveal DNA mismatch repair (MMR) activity is crucial for efficient nucleotide substitution distal from the Cas9-induced DNA break when the substitution is instructed by the 3' half of the ssODN. Furthermore, protecting the ssODN 3' end with phosphorothioate linkages enhances MMR-dependent ...[more]

PMID: 29447381

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