Hiseq 2000 2500 machines

ChIP-seq for three primary human tumors was done as described previously11 (link). Chromatin was prepared from 20 to 50 sections (25 µm each) of snap-frozen tumors obtained by microtome sectioning. The following antibodies were used: H3K4me3 (1 µg/ChIP; Diagenode, C15410003-50), H3K27me3 (1 µg/ChIP; Diagenode, C15410195), H3K4me1 (1 µg/ChIP; Diagenode, C15410194), H3K27ac (1 µg/ChIP; Diagenode, C15410196), H3K56ac (4 µl/ChIP; Active Motif, 39281), H3K9me3 (1 µg/ChIP; Diagenode, C15410193), and H3K36me3 (1 µg/ChIP; Diagenode, C15410192). Library preparation for ChIP DNA and input control DNA was performed using the NEBNext Ultra kit (New England Biolabs, E7370S/L) following the manufacturer’s instructions. Quality control for the final libraries was done by measuring the DNA concentration with the Qubit dsDNA HS assay (Life Technologies, Q32851) on Qubit 2.0 Fluorometer (Life Technologies, Q32866) and by determining library fragment sizes with the Experion DNA 1K Analysis kit (Bio-Rad, 700-7107) on the Experion Automated Electrophoresis Station (Bio-Rad, 701-7000). Libraries were sequenced on Illumina HiSeq 2000/2500 machines.

RRBS was performed as described previously51 (link),52 , starting with 100 ng of genomic DNA per sample. Custom-designed methylated and unmethylated oligonucleotides were added at a concentration of 0.1% to serve as spike-in controls for monitoring bisulfite conversion efficiency. After adaptor ligation, RRBS libraries were quantified by qPCR and pooled in combinations of six. For library enrichment, the number of PCR cycles was determined by qPCR and never exceeded 18 cycles. The library was purified twice using Agencourt AMPure XP beads (Beckman Coulter, A63880). Quality control for the final library was performed by measuring the DNA concentration with the Qubit dsDNA HS assay (ThermoFisher Scientific, Q32851) on Qubit 2.0 Fluorometer (ThermoFisher Scientific, Q32866) and by determining library fragment sizes with the Experion DNA 1K Analysis kit (Bio-Rad, 700-7107) on the Experion Automated Electrophoresis Station (Bio-Rad, 701-7000). Libraries were sequenced on Illumina HiSeq 2000/2500 machines.