Fig. 2.

Targeted analysis of DNA methylation within high-ranking DMRs by using the BisPCR² technique. (A–D) The percentage of methylation of individual CpGs within a subset of high-ranking DMRs identified in the WGBS analysis was determined using the BisPCR² technique in wild-type (n=10) and GR-mutant (n=10) adult male zebrafish brains. DMRs associated with the following genes were confirmed by BisPCR²: fkbp5 (A), foxred2 (B), lpar6a (intron 2)/ece2b (intron 8) (C), npepl1 (D); indicated on the x-axes is the number of the first nucleotide of each CpG on the respective chromosome of the Danio rerio reference genome (GRCz10 release 85). Histograms show mean percentage of methylation±s.e.m. in wild-type and GR-mutant adult brains. On average, a coverage of 4093 reads per amplicon was obtained across all samples subjected to BisPCR² analysis. Significant differences between wild-type and GR-mutant brains regarding the level of CpG methylation were determined using the Mann–Whitney test (^#P<0.06, *P<0.05,***P<0.001).