GSM1642535: Pcf11 ChIP; Saccharomyces cerevisiae; ChIP-Seq
Samples were washed in PBS and lysed by FA lysis buffer [50 mM Hepes (pH7.5), 150 mM NaCl, 1 mM EDTA, 1% Triton-X 100, 0.1% sodium deoxycholate, 0.1% SDS, protease inhibitors] with 500 µl glass beads.  Shearing of DNA was done using a Bioruptor sonicator (Diagenode) for 25 mins with 30 sec on, 30 sec off cycles.  TAP-tagged proteins were isolated from chromatin using Rabbit IgG-sepharose beads.  CL-4B-Sepharose beads were used for control IP (Sigma).  Data for H3K4me3 and RNA polymerase II was attained using ChIP-grade antibodies against H3K4me3 (Abcam, #) and RNA pol II (Abcam, #).  Both were isolated using Protein A-sepharose beads (Sigma).  IgG (Sigma, #) was used as a non-specific control for H3K4me3 and RNA polymerase II immunopreciptiations.  All samples were washed in FA lysis buffer, DOC buffer [10 mM Tris-HCl (pH8.0), 250 mM LiCl, 0.5% NP-40, 0.5% NaDOC, 1 mM EDTA] and TE buffer [10 mM Tris-HCl (pH8.0), 1 mM EDTA (pH8.0)].  Crosslinks were reversed at 65°C in TES [10 mM Tris-Cl(pH7.5), 1 mM EDTA, 1.0% SDS] and samples were treated with proteinase K.  DNA was precipitated using standard ethanol precipitation protocols and was resuspended in TE containing RNase. Libraries were prepared according to Illumina's instructions accompanying the DNA Sample Kit