Cas9_cutting_efficiency-sc-2013-04-18T15:09:03Z-2603
Cas9_cutting_efficiency
PCR products are obtained from each target loci using genomic DNA from Cas9-treated iPS cells. Subsequently, PCR products are pooled and subjected to Illumina library preparation and Miseq sequencing. This data is part of a pre-publication release. For information on the proper use of pre-publication data shared by the Wellcome Trust Sanger Institute (including details of any publication moratoria), please see http://www.sanger.ac.uk/datasharing/
The CRISPR system is becoming more popular as a tool for genome modification in mammalian cells, expecially in human induced pluripotent stem (iPS) cells and embryonic stem (ES) cells. The efficiencies of double-strand break (DSB) introduction by the CRISPR system are key for the successful engineering of the iPS/ES cell genome. To use custom Cas9 cassettes, we need to measure off-target "cutting" efficiency by a precise method, which can be adapted in a high-throughput manner. In this study, we explore the possibility of MiSeq sequencing for the "cutting" efficiency measurement. We expect that one run of MiSeq should be albe to analyze efficiency of more than 100 Cas9 cassettes.