Covalent linkage of the DNA repair template to the CRISPR-Cas9 nuclease enhances homology-directed repair

ABSTRACT: Based on the hypothesis that, enhancing the local concentration of donor oligos could increase the correction rates, we generated and tested novel CRISPR-Cas9 systems, in which the DNA repair template is covalently conjugated to Cas9 (RNPD system). To validate our results from the HEK293T reporter cells, we here tested our approach at different endogenous genomic loci and in different cell types. We first targeted the human beta globin (HBB) locus in the K562 cell line, and analyzed correction- and editing frequencies using next generation sequencing (NGS). Next we targeted the Rosa26 and proprotein convertase subtilisin/kexin type 9 (Pcsk9) locus in mouse embryonic stem cells (mESCs). Here, RNPD system was always compared to Cas9 SNAP-tag fusion proteins with uncoupled donor oligos. To al

INSTRUMENT(S): Illumina MiSeq

ORGANISM(S): Mus musculus

SUBMITTER: Helen Lindsay 

PROVIDER: E-MTAB-6808 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress