title: MOLM1_H3K4ME1
organism: Homo sapiens
description: Protocols: Cell lines were cultured using media conditions as follows. MUTZ-3: 60% alpha-MEM (Life Technologies), 20% FBS (PAA), 20% conditioned medium of cell line 5637, and 10 ng/ml hrGM-CSF (Amgen); K562, MOLM-1, and U937: 90% RPMI and 10% FBS. All cultures contained 50 U/ml penicillin and 50 μg/ml streptomycin (Sigma). JQ1 sensitivity was measured by mitochondrial dehydrogenase (MTT assay) after 6, 24, 48, and 72 h of exposure with JQ1 (5, 50, 500, or 1,000 nM) or vehicle control (DMSO, 0.05%). The cell line MOLM1 was treated for 6 hours with 1,000 nM of JQ1. Subsequently, a BRD4 Chip-Seq was performed on these cells to determine the effect of JQ1 treatment of enhancers and super enhancers. Immunoprecipitated material was de-crosslinked and was amplified using a Whole Genome Amplification kit (WGA2/WGA3; Sigma-Aldrich, Zwijndrecht, The Netherlands). Subsequently, the fragments were sheared by a Covaris shearing device (Covaris S2) and size selected by an agrose gel. ChIP experiments were performed as previously described (Bindels et al., 2012; doi: http://dx.doi.org/10.1182/blood-2011-11-393827). Immunoprecipitation of cross-linked chromatin was performed with antibodies directed against Histone H3K4me1, H3K4me3, H3K27ac, p300, and BRD4, or an equal amount of isotype IgG as background control. Three independent experiments were performed and the amount of immunoprecipitated DNA was represented relative to cross-linked input DNA.