Fig. 8
From: Successful generation of epigenetic disease model mice by targeted demethylation of the epigenome
Generation of SRS model from human cell line HEK293. a The human H19 locus is shown with CTCF-binding sites indicated by red boxes. These CTCF-binding sites have methylation-sensitive CpG sites. Location of the targets for H19-DMR and H19-promoter used for gRNAs are indicated by blue bars. Scale indicates distance in kb. b Percentages of DNA methylation by COBRA at day 5 after transfection. Error bars, mean ± s.d. (biological triplicate). c Expression analysis by qPCR. SRS model cell lines showed upregulation of H19 and downregulation of IGF2 expression compared to control cells. Error bars, mean ± s.d. (biological triplicate). *P < 0.05, **P < 0.01, ***P < 0.001 (two-tailed Student’s t test). d Anti-CTCF ChIP was performed using cells at day 9 after transfection followed by qPCR analysis. Targeting demethylation in H19-DMR increases CTCF-binding to the CTCF2, CTCF4, and P1 sites. MRPL23-AS1 was selected as negative control, and the CTCF binding site within Hox A gene cluster (hHA2) was selected as positive control. Data are normalized by hHA2. bars, mean ± s.d. (biological triplicate). *P < 0.05, n.s., not significant (one-way ANOVA)