Bisulfite Conversion Based PCR

The bisulfite reaction reaction was first described in early 1970s and was used by Frommer et al to distinguish between cytosine and 5mCytosine (5mC) in DNA. In this reaction, DNA is first treated with sodium bisulfite to convert cytosine residues to uracil in single stranded DNA, under conditions whereby 5mC remains essentially non-reactive. The DNA sequence under investigation is then amplified by PCR with primers specific for bisulfite modified DNA. Many methods based on the same principle have been developed including bisulfite genomic sequencing PCR (BSP), methylation-specific PCR (MSP), COBRA, and methylation-sensitive single nucleotide primer extension (MS-SNuPE), with the first two being the most commonly used. All methods share the same procedure of modifying DNA with sodium bisulfite as the first step and subsequently PCR amplification with primers specific for modified DNA.