GSM2773939: HGEP_2 Input (biological replicate 1); Homo sapiens; ChIP-Seq
SCC25 and HGEP cells were fixed with 1% formaldehyde for 10 min. For histone ChIP-seq we used the Chromatin Shearing Optimization Kit-Low SDS (Diagenode) with 10 million cells per cell line and 40 cycles sonication (30 sec ON, 30 sec OFF, High) in the Bioruptor Plus (Diagenode). After RNase and proteinase K incubation, chromatin shearing was confirmed as a DNA smear between 100-300 bp on a 1.5% agarose gel. Histone ChIP DNA was prepared with the Auto Histone ChIP-seq kit protein G Kit (Diagenode), with 3 μg H3K4me1, 2 μg H3K4me3, 2.4 μg H3K27ac, 2 μg H3K27me3 and 2.4 μg H3K36me3 premium polyclonal antibody (Diagenode) per 1 million cells with the SX-8G IP-Star® Compact System (Diagenode). 1% input sample for each cell line was used as a control for background experimental noise. ChIP and input DNA were quantified with the Quant-iT PicoGreen ds DNA Assay Kit (Invitrogen). ChIP-seq libraries were made with the ThruPLEXTM-FD Prep Kit (Rubicon Genomics), purified and size-selected with a 2% agarose Pippin cassette (Sage Science). Libraries were quantified with the Quant-iT PicoGreen ds DNA Assay Kit (Invitrogen) and the KAPA Library Quantification Kit (Kapa Biosystems). Quality control was performed with the Agilent Technologies 2100 Bioanalyzer. Libraries were normalized based on qPCR and pooled with the TruSeq SR Cluster Kit v3-cBot-HS (Illumina). Pooled samples were multiplexed and sequenced with the HiSeq 2500 v3 (Illumina).