Optimized sgRNA design to maximize activity and minimize off-target effects of CRISPR-Cas9 View Full Text


Ontology type: schema:ScholarlyArticle      Open Access: True


Article Info

DATE

2016-02

AUTHORS

John G Doench, Nicolo Fusi, Meagan Sullender, Mudra Hegde, Emma W Vaimberg, Katherine F Donovan, Ian Smith, Zuzana Tothova, Craig Wilen, Robert Orchard, Herbert W Virgin, Jennifer Listgarten, David E Root

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-target activity and minimize off-target effects to enable more effective and efficient genetic screens and genome engineering. More... »

PAGES

184-191

References to SciGraph publications

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  • Identifiers

    URI

    http://scigraph.springernature.com/pub.10.1038/nbt.3437

    DOI

    http://dx.doi.org/10.1038/nbt.3437

    DIMENSIONS

    https://app.dimensions.ai/details/publication/pub.1014083513

    PUBMED

    https://www.ncbi.nlm.nih.gov/pubmed/26780180


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