title: GSM4453497: Wild-type OE Cse4 input; Saccharomyces cerevisiae S288C; ChIP-Seq
library_construction_protocol: Cell suspension was distributed to screw top tubes with glass beads (0.4-0.65 mm diameter) and lysed three times for 40 seconds each, followed by a five-minute rest on ice, and lysed two times for 40 seconds each in an MP Bio FastPrep-24 5G. Lysates were collected, and the chromatin pellets  washed twice in FA lysis buffer, resuspended in FA lysis buffer, sonicated 24 times with repeated 15 seconds on/off at 20% amplitude using a Branson digital sonifer. After 3 minutes of centrifugation (13000 rpm, 4ºC), a portion of the supernatant was saved for input library constucation and the remainder incubated with anti-FLAG® M2 Affinity Gel (Sigma, A2220-5ML) overnight. The beads were washed with FA, FA-HS (500 mM NaCl), RIPA, and TE buffers twice each. The beads were resuspended in 25 mM Tris-HCl pH7.6, 100 mMNaCl, 0.5% SDS), incubated at 65°C overnight, treated with proteinase K (0.5 mg/ml) at 55°C for four hours followed by Phenol/Chloroform extraction and ethanol precipitation. Following repair using the NEB Pre-PCR Repair Mix (New England Biolabs, M0309), paired-end libraries were prepared using the NEBNext® End Prep (New England Biolabs, E7370) and NEBNext® Multiplex Oligos for Illumina (New England Biolabs, E7335).
attributes.tag: GEO Accession
attributes.value: GSM4453497