Instrument: Illumina HiSeq 3000
Strategy: ChIP-Seq
Source: GENOMIC
Selection: ChIP
Layout: SINGLE
Construction protocol: For ChIP-Seq experiments, chromatin was clarified from sonicated cell lysates by centrifugation and incubated overnight with corresponding antibodies. For RNA-Seq, cells were washed in PBS then lysed in 500 uL Trizol and total RNA was isolated using the Zymo Research Direct-zol RNA MiniPrep kit (Zymo Research #R2050) with on-column DNase digestion following the manufacturers instructions. For PRO-Seq experiments, nuclei were extracted through hypotonic lysis with a low amount of non-ionic detergent. For ChIP-Seq experiments, indexed libraries were made from precipitated DNA using the DNA Ultra II Kit (NEB) per the manufacturer's protocol. For RNA-Seq, library preperation was completed by Genewiz using their standard methods. For PRO-Seq experiments, libraries were made using RNA that was made through a biotin-run on reaction. Biotin-RNA was base hydrolyzed, and extracted. 3'-adaptors were ligated to ends and 5'caps were removed, repaired, and then 5'-adaptors were ligated as well. Purified and intact biotin-RNA containing adaptors was used in a reverase transcriptase reaction to generate cDNA and cDNA was used to amplify full library using NEB High Fidelity Phusion polymerase.