title: GSM8877193: beige_D15-rep1; Homo sapiens; OTHER
library_construction_protocol: ASCs were differentiated in 10 cm Petri dishes. Cells were washed twice in warm PBS before lysis in 4 ml Hi-ChIP buffer (10mM Tris-HCl pH 7.4, 10mM NaCl, 3 mM MgCl2; 0.01% Digitonin, 0.1% Tween 20 and 1.0% NP-40, 20 mM Na-Butyrate). Lysed cells were centrifugated at 500 g for 5 min at 4⁰C. The floating fraction (containing unlysed adipocytes) and the pellet were combined in 5 ml Hi-ChIP buffer and dounced to isolate nuclei. Nuclei were washed twice in Hi-ChIP buffer, followed by centrifugation at 500 g for 5 min at 4⁰C. The nuclei pellet was resuspended in 5 ml Nuclei buffer (10mM HEPES, 1.5 mM MgCl2, 250 mM sucrose, 0.1 % NP-40, 20 mM Na-Butyrate in PBS) and processed for Hi-ChIP according Arima-HiC+ Kit protocol (Cat Nr: A510008), with some adaptations. Briefly, 15 µg DNA equivalent of crosslinked nuclei were lysed in deionized H2O, sonicated for 10 cycles (30s on, 30s off; Bioruptor Pico). The ChIP was performed with 2.5 µg H3K27ac antibody (Diagenode C15410174). Libraries were quantified (KAPA, KK4824), amplified (KAPA, KK2620) and barcoded with xGen 2S Plus DNA library prep kit (IDT, 10009877 ) in combination with xGen 2S MID Adapter (IDT, 10009900). Libraries were sequenced to an average depth of 355 million 150 bp PE reads on an Illumina NovaSeq instrument.
identifiers.namespace: GEO
identifiers.id: GSM8877193_r1