Aberrant methylation of promoter CpG islands is known to be responsible for the alteration and silencing of genes in cancer. The data presented here describes that the CpG methylations of ERalpha and ERbeta are found in breast cancer tissues, and methylation exerts a considerable effect on gene silencing, investigated by bisulfite genomic sequencing and reverse transcriptase polymerase chain reaction. Consequently, hypermethylation of the ERalpha and ERbeta genes was found in 66.0% and 50.0% of 50 breast cancers, respectively. Eleven of 50 samples (22.0%) did not show any methylation of either ERalpha or ERbeta, whereas 19 samples (38.0%) showed methylation of both ERalpha and ERbeta. The tumors that showed aberrant methylation of ERalpha and ERbeta did not express mRNA, compared with unmethylated cases (p < 0.01). The methylated case was negatively correlated with the expression of the ERalpha protein (p < 0.01). In addition, ERbeta methylation demonstrated significant associations with the lower level of Ki67 (9.36 +/- 2.43 versus 19.68 +/- 3.42, p = 0.02). Although the number of samples was relatively small, our results suggest that DNA methylation in ERalpha and ERbeta are present in breast cancer tissue, and that the methylation causes a significant effect on transcriptional silencing. Furthermore, the CpG methylation of the ERbeta gene seems to play a role in cell proliferation of breast cancer tissue.