GSM5856260: Case1_rep2_LCL_H3K4me3; Homo sapiens; ChIP-Seq
DNA was purified by phenol-chloroform extraction and ethanol precipitation with NaOAc, and 2 µl GlycoBlue. For ChIP-seq, chromatin was crosslinked with 1 % formaldehyde at room temperature for 10 minutes. Crosslinking was quenched with 130 mM glycine, and cells were washed with cold PBS. Fixed material was stored at -80 C. Chromatin immunoprecipitation was performed using Agarose ChIP Assay Kit (Merck Millipore). Briefly, 10^7 cells were lysed by incubation on ice with lysis buffer for 15 minutes. Lysed cells were sonicated, insoluble material was removed by centrifugation and soluble material was diluted with dilution buffer. Immunoprecipitation was performed by incubation of diluted chromatin with antibodies for H3.3 (09-838, Millipore), H3K27ac (ab4729, Lot: GR276932-1, Abcam), H3K9me3 (ab8898, lot GR3244171-1, Abcam), H3K4me3 (ab8580, lot: GR273043-1, abcam), H3K4me1 ( ab8895, lot: GR283603-1, Abcam), H3K27me3  (ab6002, lot: GR275911-6-1, Abcam) and H3K36me3 (ab9050, lot: GR233723-2, Abcam) overnight. Chromatin binding to Protein A and or G/agarose slurry, washes and elution were performed according to the manufacturerâs instructions. NEBNext Ultra II DNA Library Prep kit for Illumina (New England Biolabs).