GSM7439314
SGC7901 cells were irradiated once for 400 mJ/cm2, and lysed in ice-cold wash buffer (1× PBS, 0.1% SDS, 0.5% NP-40 and 0.5% sodium deoxycholate) supplemented with a 200 U/mL RNase inhibitor (Takara) and protease inhibitor cocktail (Roche) and incubate on ice for 30 min. After centrifuge, RQ I(Promega) with a final concentration of 1 U/μL was added to the supernatant and incubate for 30 min at 37°C. Immediately afterward, a stop solution was added to the lysates to quench DNase. The mixture was then vibrated vigorously and centrifuged at 13,000 g for 20 min at 4℃ to remove cell debris. Then RNA digestion by MNase (Thermo Fisher Scientific) was performed. libraries were prepared with the KAPA RNA Hyper Prep Kit (#KK8541, KAPA) according the manufacturer's procedure. crosslinking-immunoprecipitation and high-throughput sequencing (CLIP seq)