title: GSM5508118: U2OS_masked_TH59-DB_rep1; Homo sapiens; OTHER
library_construction_protocol: To extract RNA, the input and isolated chromatin with masked TH59-DB and TH59-DB were incubated in de-crosslinking buffer (32 mM Tris-HCl pH 8.0, 320 μg/mL Protease K, 0.8% SDS) for 6 hr at 65°C. An equal volume of acidic phenol was added, then vortexed, and incubated for 5 min at room temperature. After adding an equal volume of chloroform: isoamyl alcohol (24:1, v/v) and centrifuging at 16,000 g for 5 min at 4°C, the upper aqueous phase was transferred to a new tube. This step was repeated. One-tenth volume of 3 M sodium acetate, glycogen, and double volume of ethanol were added and incubated overnight at -80°C. After centrifugation at 20,000 g for 30 min at 4ºC, the pellet was retained, washed with 70% ethanol, and dried up. The dried pellet was dissolved in diethylpyrocarbonate (DEPC)-treated water (36415-54, Nakarai) and kept at -80°C until use. RNA libraries were synthesized by the SMARTer Stranded Total RNA-Seq Kit v2 (634411, Takara).