Instrument: Illumina HiSeq 2500
Strategy: ChIP-Seq
Source: GENOMIC
Selection: ChIP
Layout: SINGLE
Construction protocol: One million cells were fixed in 1% formaldehyde/PBS buffer for 10 min at room temperature. Fixation was stopped by addition of glycine to a final concentration of 125 mM. Cells were washed 3 times with TBSE buffer. Pelleted cells were lysed in 100 µl 1% SDS lysis buffer and sonicated to 300-500bp using a Bioruptor (Diagenode). Sonicated DNA was normalized for each pair of cells with and without wildtype VHL before immuno-precipitation. The total volume of immunoprecipitation was 1 ml and the amount of antibody used was 2 µg. The input DNA was precleared with protein G Dynabeads (Life Technologies) for 1 hr at 4°C and then incubated with antibodies conjugated protein G beads overnight at 4°C. The beads were washed 3 times with cold wash buffer. Whole-genome-amplification was performed on ChIP DNA and input using the WGA4 kit (Sigma-Aldrich). The amplified DNA was used with NEBNext ChIP-Seq library prep reagent set.