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. 2020 Mar 25;21:78. doi: 10.1186/s13059-020-01989-2

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© The Author(s) 2020

Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data.

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Fig. 1 — DNA cleavage of Cas12a orthologs in vitro. a DNA cleavage activities of BfCas12a and Cas12a in vitro. The Cas12a-crRNAcomplex (100 nM) was incubated at 37 °C for 10 min with DNA substrates (see also Additional file 1: Figure S4). b Schematic of in vitro cleavage assay used to identify PAM sequence and determine global PAM specificity. c Web logo for the CeCas12a PAM. d Web logo for the BfCas12a PAM. e Normalized cleavage rates for all 4-base PAMs for CeCas12a, BfCas12a, LbCas12a, and AsCas12a. The intensity of color represented the activity of Cas12a nuclease. f Quantification of time-course in vitro cleavage reactions of Cas12a orthologs on linearized dsDNA substrates. Those were conducted at nine time points, respectively. Curves were fit using the one phase exponential decay equation