GSM3702001: H3K27ac_ChIP_Esrrb_KO_rep2; Mus musculus; ChIP-Seq
Chromatin immunoprecipitation (ChIP) was performed as previously described (Frank et al., 2001) with minor modifications outlined below. Chromatin was fixed with 1% Formaldehyde (Sigma-Aldrich) for 10 minutes and sonicated on a bioruptor (Diagenode) to produce fragments of 100-500 bp, and ChIPs performed with Protein G-coupled magnetic Dynabeads (Invitrogen) and the following antibodies : 8 µg MED1 (A300-793A Bethyl Laboratories). The amounts of chromatin (protein) used in each ChIP were as follows: 800 µg (Med1).  Following washes of bound DNA-protein complexes, DNA was eluted in 1% SDS and treated with 40 ng/µl RNaseA following 0.2 µg/µl Proteinase K. After phenol/chloroform purification, DNA was then precipitated at -20°C with 20-30 μg GlycoBlue carrier (Invitrogen), 1/10 volumes of 3 M NaAc and 2 volumes of 100% ethanol. Sequencing libraries were prepared using the NEBNext® UltraTM DNA Library Prep Kit and Multiplex Oligos (New England Biolabs) from 5 ng of DNA. Following analysis on an Agilent Bioanalyzer, libraries were pooled and sequenced on an Illumina NextSeq500.