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bioRxiv · 10.1101/057802

Genome-wide specificity profiles of CRISPR-Cas Cpf1 nucleases in human cells

Benjamin P Kleinstiver·Shengdar Q Tsai·Michelle S Prew·Nhu T Nguyen·Moira M Welch·Jose M Lopez·Zachary R McCaw·Martin J Aryee·J. Keith Joung
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Abstract

CRISPR-Cas Cpf1 nucleases have recently been described as an alternative genome-editing platform1, yet their activities and genome-wide specificities remain largely undefined. Here we show that two Cpf1 nucleases function robustly in human cells with on-target efficiencies comparable to those of the widely used Streptococcus pyogenes Cas9 (SpCas9)2-5. We also demonstrate that four to six bases at the 3 end of the short CRISPR RNA (crRNA) used to program Cpf1 are insensitive to single base mismatches but that many of the other bases within the crRNA targeting region are highly sensitive to single or double substitutions. Consistent with these results, GUIDE-seq and targeted deep sequencing analyses of two Cpf1 nucleases revealed no detectable off-target cleavage for over half of 20 different crRNAs we examined. Our results suggest that the two Cpf1 nucleases we characterized gen ...

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Benjamin P Kleinstiver, Shengdar Q Tsai, Michelle S Prew, Nhu T Nguyen, Moira M Welch, Jose M Lopez, Zachary R McCaw, Martin J Aryee, J. Keith Joung. 2016-06-09. Genome-wide specificity profiles of CRISPR-Cas Cpf1 nucleases in human cells. https://doi.org/10.1101/057802

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