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. 2021 Oct 22;12(11):981. doi: 10.1038/s41419-021-04292-5

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© The Author(s) 2021

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. 4 — A Left, RT-qPCR analysis of endoderm genes FOXA2, SOX17 and GATA6; Right, RT-qPCR analysis of CHD8 and pluripotent genes OCT4 and SOX2 at day 3 of differentiation (n = 5). Wildtype ESCs at day 0 served as control. B Immunofluorescence of endoderm marker FOXA2 and SOX17 in WT and CHD8KO groups at day 3 of differentiation. The percentage of FOXA2- or SOX17-positive cells was calculated (n = 4). Scale bar = 200 µm. C Flow cytometric analysis showing the percentage of SOX17-positive cells in differentiated WT and CHD8-KO cells (n = 5). D, E AnnexinV/PI assay (D) and Cell cycle analysis using PI staining (E) in differentiated WT and CHD8-KO definitive endoderm cells (n = 5). The populations of different cell cycle phases were calculated in FlowJo (v10.4.0). The error bars represented the mean ± SD and the significance level was calculated by Student’s t-test (two-tailed, equal variance) (ns means not statistically significant, *P < 0.05, **P < 0.01, ***P < 0.001).