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. 2021 Nov 25;4:1334. doi: 10.1038/s42003-021-02849-w

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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 The temporal changes in methylation of promoter regions were analyzed by bisulfite sequencing. Granulosa cells were collected from secondary follicles with multilayered granulosa cells or antral follicles (3-week mice not treated with eCG) before or after eCG injection. Values are represented as the mean ± SEM (n = 3 biological replicates). Significant differences were observed between SF and eCG at 0 h (p < 0.05). *Significant differences were induced by eCG injection compared with before eCG injection (0 h) (p < 0.05). SF, secondary follicles. b, c The kinetic changes of the opened chromatin region (OCR) detected by FAIRE-qPCR (b) or the acetylated H3K27 detected by ChIP assay (c) in the promoter region of each gene in granulosa cells of secondary follicles with multilayered granulosa cells or antral follicles (3-week mice not treated with eCG) before or after eCG injection. The SF value was set as 1 and the data are expressed as fold induction. Values are represented as the mean ± SEM (n = 3 biological replicates). Significant differences in percentage values were transformed into normally distributed numbers by angle transformation and then analyzed by one-way ANOVA. Tukey–Kramer was used as post hoc test. *Significant differences were induced by eCG injection compared with before eCG injection (0 h) (p < 0.05). d Kinetic changes of the expression level of each gene in granulosa cells. Levels of mRNA were normalized to that of L19. Values are represented as the mean ± SEM (n = 3 biological replicates). Significant differences were observed between SF and eCG at 0 h (p < 0.05). *Significant differences were induced by eCG injection compared with before eCG injection (0 h) (p < 0.05).