GSM3293868: WT_polII_ChIPSeq_rep2; Saccharomyces cerevisiae; ChIP-Seq
Yeast samples were fixed with 1.035% formaldehyde for 15 minutes and neutralized with 0.135 M glycine for 5 minutes.  Washed cells were lysed in FA-lysis buffer (50 mM HEPES-KOH, pH 7.9; 150 mM NaCl; 1% Triton X-100; 1 mM EDTA; 0.1% sodium deoxycholate; 0.1% SDS) with grinding in liquid nitrogen..The resulting 4 ml chromatin were incubated with 40 l 4H8 antibody and 200 l pre-washed protein G beads for 5 hours at 4°C.  The beads were washed as described above and were then eluted in two steps: 1) in 150 l TE + 1% SDS at 65°C for 15 minutes, and 2) in 150 l TE + 0.67% SDS at 65°C for 10 minutes.  Samples were de-crosslinked overnight at 65°C and then treated with Proteinase K for 2-4 hours at 42°C.  LiCl (to 0.4 M), 20 g glycogen and 1 ml 100% ethanol were added to precipitate the DNA.  The DNA pellets were washed in 70% ethanol and were further purified using MinElute columns (Qiagen).  The eluted DNA was submitted to the Tufts Genomics Core Facility for TruSeq ChIP library preparation and for 50 nt single-end sequencing on the Illumina HiSeq 2500 system. Illumina TruSeq ChIP library preparation
GEO Accession
GSM3293868