Bisulfite Sequencing

Bisulfite sequencing is a widely used method for mapping DNA methylation patterns at single-nucleotide resolution. DNA methylation, primarily the addition of a methyl group to cytosine residues, is the first epigenetic modification ever discovered and remains one of the most extensively studied. Changes in cytosine methylation play key roles in the regulation of gene expression, genome stability, and cellular differentiation. These methylation patterns can also be inherited across cell divisions and, in some cases, across generations, providing a mechanistic basis for epigenetic inheritance.

The core principle of bisulfite sequencing relies on the chemical conversion of unmethylated cytosines. When genomic DNA is treated with bisulfite, unmethylated cytosines are deaminated and converted into uracil, while 5-methylcytosines remain unchanged. During subsequent PCR amplification and sequencing, uracils are read as thymines. As a result, methylated cytosines appear as cytosines in the sequencing data, whereas unmethylated cytosines appear as thymines. By comparing the treated DNA sequence to the original reference, researchers can infer the methylation status of each cytosine with single-base precision.


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