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. 2022 Mar 17;13:1425. doi: 10.1038/s41467-022-29089-8

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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 — a The schematic of puromycin depletion assay. b, c The PAM results of SpCas9 and FrCas9 from the puromycin depletion assays. Data are presented as mean values ± S.E.M. (n = 3 biological independent replicates). d The TIDE assay showed FrCas9-induced indel rates in 3 human genes by 12 sgRNAs, which differed in the 2^nd PAM base. e Genome editing by FrCas9 in the human RNF2 gene, validated by double-stranded oligodeoxynucleotide (dsODN) breakpoint PCR. And the Sanger sequencing was in Supplementary Fig. 2. Uncropped gel image is provided in Source Data. f The schematic representation of the sgRNA:target DNA complex. g The efficiency of sgRNAs with truncated scaffolds of FrCas9 assayed by target amplicon sequencing. Data are presented as mean values ± S.E.M. (n = 3 biological independent replicates). h The FrCas9 editing efficiencies with 19–23 bp spacer lengths in three human sites by target amplicon sequencing. i The FrCas9 editing efficiencies with 21 and 22 bp spacer lengths in five human sites by TIDE (n = 3 biological independent replicates). Source data are provided with this paper.