GSM4826146: WT_Sua7-FLAG_rep2; Saccharomyces cerevisiae; ChIP-Seq
Cells were crosslinked as described in (Brown, Howe et al., 2018). Crosslinked cells were resuspended in cold FA‐150 buffer (10 mM HEPES pH 7.9, 150 mM NaCl, 0.1% SDS, 0.1% sodium deoxycholate, 1% Triton X‐100) and mixed with pre-crosslinked with S.pombe in 5:1 ratio (final S.pombe percentage being 16.7 %). The cell suspension was lysed with glass beads using the MagnaLyser (Roche; 6 x 45 s runs, 2500 g, 4°C). The lysate was sheared 30-40 min with a bioruptor sonicator 30 sec ON/30 sec OFF at high setting. The sheared lysate was cleared by centrifuge (10,000 g, 15 min, 4°C) and the supernatant was incubated with anti-FLAG (M2) antibody for 15–20 h rotating at 4°C. Bound chromatin was immunoprecipitated for 90 min at 22°C with pre‐blocked Dynabeads protein G. Beads and attached chromatin were pelleted, washed, and immunoprecipitated chromatin was eluted from the beads as described in (Brown, Howe et al., 2018). DNA was eluted with Zymoclean ChIP concentrator kit according to the manufacturer's instructions. standard NEBNext Ultra II DNA Library Prep Kit for Illumina