RRBS of Homo sapiens : whole blood
To construct the MSP1 and Apek1 digested reduced-representation bisulfite sequencing (RRBS) library, 500 ng of input genomic DNA was assembled into 50 l of reactions with MspI (NEB), incubated at 37C for 24-26 h. ApeKI (NEB) was then added and incubated at 75C for 1620 h. The digested products were purified with a MiniElute PCR Purification Kit (Qiagen). After purification, the digested products were blunt-ended, and then dA was added, followed by methylated-adapter ligation. A range of 160-420 adapter-ligated fraction was excised from a 14% agarose gel. Bisulfite conversion was conducted using a ZYMO EZ DNA Methylation-Gold Kit (ZYMO), following the manufacturers instructions. The final libraries were generated by PCR amplification using PfuTurbo Cx Hotstart DNA polymerase (Agilent technologies, Santa Clara, CA, USA).