GSM1541741: USV101 tagged capsule inducing mock; Cryptococcus neoformans; ChIP-Seq
The cells were fixed with formaldehyde, lysed by mechanical bead-beating, and the cell debris was  removed by centrifugation. The supernatant fraction, containing isolated chromatin, was sheared by sonication, cleared again by centrifugation, and an aliquot was reserved as ‘input’. The remaining material was incubated with rabbit IgG anti-HA antibody (Abcam) tethered to protein A sepharose (‘IP’) or sepharose alone (‘mock’) overnight at 4°C. The beads were then washed, incubated at 65°C to reverse DNA-DNA and DNA-protein crosslinks, and the DNA recovered by phenol/chloroform/isoamyl alcohol (25:24:1) extraction, ethanol precipitation, and resuspension in nuclease-free water. ChIP-DNA for for all samples was end-repaired with Klenow DNA Polymerase and the DNA was purified with AMPure XP System beads (Beckman Coulter Genomics) and modified with A-tails using Klenow exo- before ligation to adapters to incorporate 7-base index sequences using T4 DNA ligase (Enzymatics). Adapter addition was confirmed on an Agilent 2100 bioanalyzer, and the DNA was PCR-amplified and then gel purified to remove adapter dimers and select sizes optimal for high-throughput sequencing (150 to 300 bp). Samples were submitted to the Washington University Genome Technology Access Center for library preparation and multiplex sequencing.