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. 2020 Oct 20;10:17753. doi: 10.1038/s41598-020-74772-9

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© The Author(s) 2020

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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Screening of gRNAs targeting B2M or α chains of HLA-II genes. Raji cells were transfected with the indicated gRNAs complexed with Cas9 protein (gRNA/Cas9) and cultured for 7 days. (A) Target deletion efficiency of the indicated gRNAs as detected by flow cytometry in Raji cells. Percentages of target HLA-negative cells out of total cells are presented as % HLA neg (n = 3). Error bars indicate mean ± standard error of the mean (SEM). (B) Amount of HLA-targeted gene disruption measured by a mismatch-sensitive enzyme assay on DNA amplified from target-gRNA/Cas9- or non-targeting gRNA/Cas9-transfected Raji cells. Expected sizes of the DNA fragments are specified in separate tables below. Representative images are shown from three independent experiments. Full-length gels are presented in Supplementary Figure S3. M, DNA Marker. (C) On-target and off-target mutagenesis measured by NGS on genomic DNA amplified from indicated gRNA/Cas9-transfected Raji cells (n = 3). Error bars indicate mean ± SEM. On, on-target. Off, off-target.