Sequencing based methylation analysis

Methylated cytosines are insensitive to conversion to thymine (uracil) by bisulfite. Treating DNA with bisulfite before sequencing with high throughput technology will reveal the position of methylated cytosines, and their degree of methylation by read-counting.

Whole genome sequencing doesn’t require prior knowledge of methylation signals, but requires to sequence the whole genome at high coverage.

Reduced Representation Bisulfite Sequencing is a way to avoid whole genome sequencing and reduce sequencing cost. It is based on the use of the MspI restriction enzyme that preferentially cuts in CpG islands. Small DNA fragments after MspI digestion of genomic DNA will manly come from CpG islands. By only sequencing these smaller fragment, CpG islands will preferentially be targeted.