Breast cancer cells lose methylation of E-cadherin promoter methylation following hepatocyte coculture. A) Nested PCR method to detect methylation status of the E-cadherin promoter in a six day time course of hepatocye coculture. Above, bisulfite-treated DNA is amplified with primers that exclude CpG islands to amplify a 270 bp region independent of methylation status. Below, nested primers anneal to the 270 bp target to amplify a methylated (112 bp) or unmethylated (120 bp) fragment in the six day time course. MCF7 is used an unmethylated control. B) Bisulfite sequencing of CpG islands in the E-cadherin promoter. Figure adapted from Corn et al . CpG islands are indicated as vertical lines on map; each CpG island is represented a circle. MCF7, MDA-MB-231, and MDA-MB-435 were sequenced on days 1,3, and 5 coculture. Open circle, unmethylated CpG; closed circle, methylated CpG; filled circle, mixed quality values.