GSM4851971: WT_mock_rep1; Homo sapiens; ChIP-Seq
ChIP-seq: Chromatin was sonicated to approximately 1 nucleosome using a Bioruptor waterbath sonicator (Diagenode). Chromatin of ~5x106 cells was incubated with 3ug antibody (RNAPII, rabbit polyclonal, Bethyl laboratories, A304-405A; PAF1, rabbit polyclonal, Bethyl laboratories, A300-172A) over night at 4°C, followed by a 1.5 hrs protein-chromatin pull-down with a 1:1 mix of protein A and protein G Dynabeads (Thermo Fisher; 10001D and 10003D). ChIP samples were washed extensively, followed by decrosslinking for 4 hrs at 65°C in the presence of proteinase K. DNA was purified using a Qiagen MinElute kit. Sample libraries were prepared using Hifi Kapa sample prep kit and A-T mediated ligation of Nextflex or xGen UDI-UMI adapters. Samples were sequenced using an Illumina NextSeq500 or HiSeqX, using paired-end sequencing with 42 bp or 151 bp from each end. BrU-seq: cDNA libraries were made from the Bru-labeled RNA using the Illumina TruSeq library kit and paired-end 151 bp sequenced using the Illumina NovaSeq platform at the University of Michigan DNA Sequencing Core. Only single-end sequencing data was used for downstream analyses