Studying DNA Methylation Genome-Wide by Bisulfite Sequencing from Low Amounts of DNA in Mammals

Abstract

DNA methylation is extensively reprogrammed during mammalian embryogenesis and germ cell development. Protocols for genome-wide bisulfite sequencing enable the quantification of DNA methylation with high precision and single base-pair resolution; however they can be limited by the necessity for high amounts of DNA. Here we describe optimized reduced representation bisulfite sequencing (RRBS) and whole genome bisulfite sequencing (WGBS) protocols for low amounts of DNA, which include steps to estimate the minimal number of PCR cycles needed for the final library preparation to minimize PCR biases. These protocols require no more than 5 ng DNA and can easily be applied to mammalian cells available in small quantities such as early embryos or primordial germ cells.

Keywords: Bisulfite conversion; DNA methylation; DNA methylome; Epigenome; RRBS; Reduced representation bisulfite sequencing; WGBS; Whole genome bisulfite sequencing.