GSM5235631: input of Swc3-ΔZ yeast; Saccharomyces cerevisiae; ChIP-Seq
Yeast cells were fixed with 1% formaldehyde for 30 min at room temperature, followed by quenching fixation with 0.125 M glycine for 5 min. Cell wall was disrupted using Ziconia beads and lysed in lysis buffer (50 mM Tris pH7.5, 1mM EDTA pH8.0, 1% Triton X-100, 150 mM NaCl, 1×EDTA-free protease inhibitor cocktail). Chromatin was sonicated to an average length of approximately 250 bp. The lysates were clarified by centrifugation, and the supernatants were incubated with protein A/G beads overnight at 4 °C to remove non-specific binding. Supernatant were collected by centrifugation and added anti-Htz1 antibody for incubation 4 h. The Antibody-Chromatin complex were pulled down with protein A Dynabeads and were washed twice with TBST buffer, twice with TE buffer, followed by incubation with TES buffer (50 mM Tris pH7.5, 10 mM EDTA pH8.0, 1% SDS) to elute the complexes. Following cross-link reversal, Rnase A and Proteinase K treatment, phenol-chloroform extraction and ethanol precipitation DNA samples were used to prepared libraries with Kapa hyper prep kit and NEB Next multiples oligos for Illumina (NEB E7335) according to the operation instructions.