title: GSM5107575: ChIP_14: Healthy donor 06 mDC H3K27ac; Homo sapiens; OTHER
library_construction_protocol: Briefly, isolated mDCs from ECs and HIVNs were bound to concanavalin A-coated magnetic beads (Bangs Laboratories), followed by cell permeabilization using 0.01% digitonin (Millipore) in wash buffer. Primary antibodies against intracellular H3K27ac, H3K27me3 and H3K4me3 histone marks (Cell Signaling Technology) were added and cells were incubated at 4oC for overnight. Guinea pig anti-rabbit IgG antibody (Antibodies-online) was used as a negative control. The next day, fusion protein CUTANA pA/G-MNase (20x) (Epicypher) was added and cells were incubated at 4oC for 1 hour, followed by chromatin digestion using 100mM CaCl2 (Fisher Scientific). Stop buffer was then added to stop the chromatin digestion. DNA fragments were extracted using phenol chloroform (Invitrogen) and the extracted DNA was eluted in 1mM Tris-HCl pH 8.0 (Fisher Scientific) + 0.1 mM EDTA (Sigma Aldrich). The DNA library was prepared using NEBNext Ultra II DNA Library Prep Kit for Illumina Sequencing (New England Biolabs).