GSM9313030: VCaP_100nM_DALTAC_1_3h_FOXA1; Homo sapiens; ChIP-Seq
In brief, cells were crosslinked in 1% formaldehyde, quenched with glycine, and lysed for chromatin extraction. Chromatin was sheared to 200–600 bp fragments using a Bioruptor (Diagenode) and subjected to immunoprecipitation with antibodies against transcription factors (AR, FOXA1, p300, ERG and BRD4) or histone marks (H3K27ac, H2BK20ac, and RNA Pol II) using the iDeal ChIP-seq Kit (Diagenode). Drosophila spike-in chromatin (Active Motif, 53083) and spike-in antibody (Active Motif, 61686) were added during the overnight incubation. After reverse crosslinking and DNA purification, sequencing libraries were prepared and sequenced on an Illumina HiSeq 2500 platform. ChIP samples (1–10 ng) were converted to blunt-ended fragments using T4 DNA polymerase, Escherichia coli DNA polymerase I large fragment (Klenow polymerase), and T4 polynucleotide kinase (New England BioLabs (NEB)). A single adenine base was added to fragment ends by Klenow fragment (3′ to 5′ exo minus; NEB), followed by ligation of Illumina adaptors (Quick ligase, NEB). The adaptor-ligated DNA fragments were enriched by PCR using the Illumina Barcode primers and Phusion DNA polymerase (NEB). PCR products were size-selected using 3% NuSieve agarose gels (Lonza) followed by gel extraction using QIAEX II reagents (Qiagen). Libraries were quantified and quality checked using the Bioanalyzer 2100 (Agilent) and sequenced on the Illumina HiSeq 2500 Sequencer (125-nucleotide read length).