Ez dna methylation lightning kit

Placenta samples were frozen immediately after birth. As previously described [20 (link)], 5-μg placental DNA was extracted, sonicated to ∼300 bp, and methylated Illumina adapters were ligated to the ends using NEB’s NEBNext DNA library prep kit. The library was bisulfite converted using Zymo’s EZ DNA Methylation-Lightning Kit, amplified for 14 cycles using PfuTurbo Cx, purified with Agencourt AMPure XP beads, and sequenced on an Illumina HiSeq 2000. Laboratory personnel were blinded to the case status of the samples, and samples were arranged randomly. Reads were mapped to the hg19 version of the human genome using BS Seeker [50 (link)]. To prevent clonal PCR amplification biases, only one read out of those with identical genomic positions was kept. CpG site methylation data were combined from both DNA strands. Sequencing information for each placenta sample, including the total reads, total mapped reads, coverage, and conversion efficiency is provided in Table S9.