Chromatin immunoprecipitation (ChIP)

ChIP assays were carried out according to the protocol described previously^24,45. The average fragment size of soluble chromatin fragments after sonication was ∼500 bp. In vivo cross-linking, chromatin purification and immunoprecipitations were as described previously⁴⁶. A total of 150 µg of chromatin was immunoprecipitated using 2 µg of anti-γH2AX or without antibody (mock). The other antibodies used were obtained from various commercial suppliers mentioned in antibodies section. For ChIP-qPCR, immunoprecipitated and input DNA were analyzed in triplicate by real time quantitative-PCR (primer sequences as mentioned previously²⁴). Real-time PCR was performed using Applied Biosystems® SYBR® Green PCR Master Mix on a Stratagene® MX3005 P instrument, and quantitative reverse transcription-PCR (RT-PCR) data were analyzed by using the 2ΔΔ^−CT method as described previously⁴⁷.