GSM8009059: E233S,Input; Saccharolobus islandicus; ChIP-Seq
Formaldehyde was added to an exponentially growing culture at the final concentration of 1 % and the culture was incubated on a shaking platform for 15 min at room temperature for cross-linking. The whole cells were harvested by centrifugation at 5,000 g for 10 min and washed with PBS buffer. The pellet was resuspended in 3 mL TBSTT buffer (20 mM Tris, 150 mM NaCl, 0.1 % Tween-20, 0.1 % Triton X-100, pH 7.5) and sonicated on ice by JY92- Ⅱ Ultrasonicator (Scientz biotechnology, Ningbo, China) to obtained the DNA fragment with a size around 200-1000 bp. Immunoprecipitation was performed by incubating 5 µL anti-PerR antibody-coated protein A beads (Cytiva, USA) in TBSTT buffer. ChIP-seq libraries were constructed using NEBNext DNA library prep kit per manufacturer's instructions. 50ng of DNA was ligated to adaptor with NEBNext DNA library prep kit, after end repair and adenylation. Size selection and gel purification, followed by PCR with primers specific to the sequencing platform. Library concentration was quantified by Qubit® 2.0 Flurometer (Life Technologies, CA, USA) and quantitative PCR, and the insert size was assayed on Agilent Bioanalyzer 2100 system.