ATAC-seq of human dermal papilla cells in osteogenic media replicate 2
Cells were twice in ice-cold PBS and incubated with 0.5% trypsin-EDTA for 5 minutes until the cells dissociated from the dish.  Media was then added to the cells, which were centrifuged at 300g for 4 minutes to form a pellet.  The pellet was re-suspended in fresh media, counted, and the volume of media adjusted to attain a 4.5 x 104 cells/ml concentration.  1 ml of each suspension was then transferred to a 1.5 ml Eppendorf tube, centrifuged at 500g for 5 minutes at 4C, and re-suspended in 50l of ice-cold PBS.  Following a further centrifugation under the same conditions, the pellet was re-suspended in 50l of a transposition mix containing TD Buffer (25l), TDE1 (2.5l), nuclease free water (22l) and digitonin 1% (0.5l).  Samples were incubated with the transposition mix for 30 minutes, undergoing a brief vortex every 10 minutes.  Immediately after incubation, transposed samples were purified using the MinElute PCR Purification kit (Qiagen, 28004) as per the manufacturers instructions and PCR amplified for 11-13 cycles.  Right side size SPRI bead selection with a ratio of x0.5 was then performed to remove fragments under 100 and over 1000bp in size.  Quality control was performed using Qubit High Sensitivity DNA assay and Agilent Technologies Bioanalyzer, to check for nucleosome banding within the generated libraries.  Libraries were sequenced using 75bp paired-end-reads on 2 lanes of an Illumina HiSeq 4000 instrument.