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. 2023 Jan 4;613(7944):588–594. doi: 10.1038/s41586-022-05559-3

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© The Author(s) 2023

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 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/.

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Extended Data Fig. 3 — a, Diagram of the predicted secondary structure of the SuCas12a2 direct repeat. Cleavage sites of SuCas12a2 and Cas12a are indicated. b, Sequencing coverage of the cDNA mapped to the crRNA locus in E. coli BL21(AI) expressing SuCas12a2. Coverage for all of the quality-filtered reads above 10 nts as well as reads between 10 and 55 nts mapped to the plus strand are shown. c, In vitro processing of a 56-nt pre-crRNA incubated for 60 min in the presence of apo-SuCas12a2 or two mutants predicted to disrupt crRNA processing. d, In vitro SuCas12a2-mediated cleavage of a pre-crRNA containing three direct repeats and three spacers (3× crRNA). Reactions containing apo-SuCas12a2 are indicated. Time points after mixing the cleavage reaction with apo-SuCas12a2 are indicated. The estimated sizes of the pre-crRNA and crRNAs after cleavage are indicated on the left. The last two lanes contain RNA extracted from SuCas12a2 co-expressed with a CRISPR array. The difference in size between the major crRNA band in the in vitro assay (~ 67 nt) and the crRNA extracted from SuCas12a2 bound to E. coli-expressed crRNA (~42) may be due to further trimming by 3′ exonucleases in the cell. For gel source data, see Supplementary Fig. 1.