In mammals, 5-methylcytosine involves the covalent attachment of a methyl group to the 5’ carbon position of cytosine, conferring an additional ability of signaling and regulatory function. As one of the major epigenetic modifications, 5-methylcytosine plays a significant role in many biological processes, including gene silencing, suppression of transposable and repetitive sequences, genomic imprinting, X chromosome inactivation. Detection and quantification of methylation are critical to understand gene expression and other processes subjected to epigenetic regulation.
Whole genome bisulfite sequencing is the gold-standard approach to acquiring comprehensive base-pair resolution and quantitative information at most genomic methylated cytosines, allowing for unbiased genome-wide DNA methylation profiling. In the whole genome bisulfite sequencing process, the sodium bisulfite treatment of genomic DNA can convert unmethylated cytosines into uracil, while methylated cytosines keep intact. Following this step, PCR amplification, library preparation, and the next-generation sequencing are performed. Finally, through comparisons between untreated and sodium bisulfite-treated sequences, it’s viable to determine which nucleotide sites are methylated.