DNA Bisulfite Sequencing for Single-Nucleotide-Resolution DNA Methylation Detection

Abstract

DNA methylation plays an important role in multiple biological processes. Therefore, methodologies that can detect changes in DNA methylation are of general importance. A popular and reliable method for measuring DNA methylation status is DNA bisulfite sequencing. This protocol details the steps required for bisulfite conversion and analysis of either genes or a specific genomic region. Denatured DNA (i.e., single-stranded DNA) is treated with sodium bisulfite under conditions that preferentially convert unmethylated cytosine (C) residues to uracil (U) residues while methylated cytosines remain unmodified. The converted DNA can then be amplified using a gene-specific primer. U is amplified as thymidine (T) in polymerase chain reaction (PCR) and detected as T during DNA sequencing.