GSM7017075: MCF10A-ev-r1-H3K27ac; Homo sapiens; OTHER
MinElute Kit (Qiagen, #28004) To generate sequencing libraries, real-time PCR was performed in a total volume of 50 µl using 25 µl NEBNext High Fidelity 2x Mix (New England Biolabs, #M0541), 0.5 µl 100x SYBR Green, 19.5 µl tagmented eluate, and 5 µl of Nextera UDI primer (replicates R1, R3, and R4) or 2.5 µl primer T5McP1n and 2.5 µl barcode primer (replicate R2) for each sample with the following program: 72°C, 5 minutes (gap repair); 98 °C, 30 seconds (initial melting); 98°C, 10 seconds; 63°C, 10 seconds; 72°C, 10 seconds (cycling). The reaction was stopped after an increase of five or more fluorescence units. Libraries were purified using AMPure XP beads with a bead-to-DBA ratio of 1.4:1 and 12 µl elution buffer for single-phase purification (anti-HA antibodies and IgG) and with bead-to-DBA ratios of 0.5:1 and 1.4:1 for biphasic purification (anti-H3K27me3 and anti-H3K27ac). ACT-Seq, paired-end