Illumina DNA Methylation Profiling

Genomic DNA was extracted from buffy coat fractions of anti-coagulated blood samples using Chemagic DNA buffy coat kit (PerkinElmer, Germany) with Chemagic Magnatic Separation Module 1 and Chemagic Prime 8 Automated Workstation, and was subsequently bisulfite converted using the EZ-96DNA Methylation-Lightning^TMMagPrep from Zymo according to the manufacturer’s instructions. DNA methylation levels were measured on Illumina iScan using Illumina’s Human MethylationEPIC BeadChip. The methylation level for each probe was derived as a beta value representing the fractional level of DNA methylation at that probe. Sample-level and probe-level quality control was performed using the ‘minfi’ package [38 (link)] in R (version 3.5.0). Samples with sex mismatch or a missing rate at > 1% across all probes were excluded. Probes with a missing rate > 1% (at a detection p value > 0.01) were also excluded following previously published recommendation guidelines for analyzing methylation data [39 (link)].