DNA methylation analysis based on bisulfite conversion

Exposing DNA to bisulfite rapidly leads to the deamination of unmethylated cytosines which are converted to 6-sulfonyluracil. At high pH, 6-sulfonyluracil is desulfonated to uracil, which ultimately after amplification will translate into thymidine, while methylated cytosines will not be converted. Comparing this converted DNA to the original unconverted sequence enables detailed evaluation of the location and abundance of methylated sites in CpG islands. High-resolution melting (HRM) analysis provides a rapid screening tool to accurately detect changes in CpG methylation status of bisulfite converted DNA. Detailed quantitative measures of percent methylation at individual CpG sites are obtained by Pyrosequencing. Alternative methods for broad-scale methylation analysis include methylation-specific PCR (MSP) which is highly specific and sensitive. Commercial kits are available for all forms of analysis.