GSM8207955: Brn1-HA ChIP, ΔSgo1 no tension, Input; Saccharomyces cerevisiae; ChIP-Seq
Cells were lysed by beating in MP biomedicals FastPrep-24 machine for 30 seconds three times with 10 minute intervals on ice. Cell pellets were washed once in FA lysis buffer/0.1% SDS before sonicating in a Bioruptor (Diagenode) for two times 30 cycles at 30sec on/off intervals at "High" setting. The supernatant was subject to IP with Roche 12CA5 anti-HA antibody conjugated to Dynabeads over night. Dynabeads were washed and bound protein eluted using TES buffer. Samples were decrosslinked with Proteinase K and purified using Promega Wizard kit. DNA was blunted using NEB Quick Blunting kit followed by Ampure purification of fragments over 100bp. dA tails were added using Klenow (exo-) and NextFlex adaptors ligated to each sample. This was followed by two Ampure purifications of fragments over 100bp and then fragments over 150-200bp. Libraries were amplified using NextFlex primers and Phusion DNA polymerase. Lastly, double-sided Ampure purification was carried out to obtain fragments between 100-300bp in size.