Optimized sgRNA design to maximize activity and minimize off-target effects of CRISPR-Cas9.

ABSTRACT: CRISPR-Cas9-based genetic screens are a powerful new tool in biology. By simply altering the sequence of the single-guide RNA (sgRNA), one can reprogram Cas9 to target different sites in the genome with relative ease, but the on-target activity and off-target effects of individual sgRNAs can vary widely. Here, we use recently devised sgRNA design rules to create human and mouse genome-wide libraries, perform positive and negative selection screens and observe that the use of these rules produced improved results. Additionally, we profile the off-target activity of thousands of sgRNAs and develop a metric to predict off-target sites. We incorporate these findings from large-scale, empirical data to improve our computational design rules and create optimized sgRNA libraries that maximize on-

SUBMITTER: Doench JG 

PROVIDER: S-EPMC4744125 | biostudies-literature | 2016 Feb

REPOSITORIES: biostudies-literature