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. 2022 Aug 30;13:4917. doi: 10.1038/s41467-022-32618-0

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© The Author(s) 2022

Open Access This 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 license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license 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 license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 2 — Screening of all 64 possible target sites containing each of the three-nucleotide PAM sequences for trans cleavage activity by EcoCas3 (a) and by LbaCas12a (b). The heat maps represent the RFU per min for collateral cleavage activity. c Trans ssDNA cleavage by a crRNA-complementary or non-complementary ssDNA (TS or NTS, respectively). EcoCas3/EcoCascade partially activated by TS ssDNA in a PAM-dependent manner (3′-TTC-5′ only). d LbaCas12a activated by TS ssDNA in a PAM-independent manner (both 3′-AAAC-5′ and 3′-TTTG-5′). e Screening of all 64 possible target sites containing each of the three-nucleotide PAM sequences for collateral cleavage activity by the TS ssDNA. f Collateral cleavage activated by crRNA-complementary ssDNA, where the PAM region was double stranded but the rest of the protospacer was single stranded. dsDNA-activated collateral cleavage is also shown to the right. Row data in the heatmaps of a, b, e is available in the Source data file. Data in c, f are presented for n = 3 independent measurements and mean value, error bars represent SD values. Data in d are presented for n = 2 independent experiments with central value.