Principle of DNA methylation analysis by bisulfite treatment. On a DNA sequence, unmethylated cytosines are converted to uracil by the bisulfite treatment and to thymines after PCR amplification. After sequencing, the level of methylation is detected by counting cytosines and thymines for each position. m, methyl group on cytosine; OT, Original Top strand; CTOT, strand complementary to the original top strand; OB, original bottom strand; CTOB, strand complementary to the original bottom strand. Available via license: CC BY 4.0 (27).