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.
journal_name
Nat Biotechnoljournal_title
Nature biotechnologyauthors
Doench JG,Fusi N,Sullender M,Hegde M,Vaimberg EW,Donovan KF,Smith I,Tothova Z,Wilen C,Orchard R,Virgin HW,Listgarten J,Root DEdoi
10.1038/nbt.3437subject
Has Abstractpub_date
2016-02-01 00:00:00pages
184-191issue
2eissn
1087-0156issn
1546-1696journal_volume
34pub_type
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