GSM4395590: TSC346_BJ_ATAC_shSMAD3-72; Homo sapiens; ATAC-seq
~50,000 BJ fibroblasts (senescent or quiescent via contact inhibition) were collected via trypsinization, washed in PBS on ice, resuspended in 200uL ice cold ATAC-seq lysis buffer (10mM Tris-HCl pH 7.5, 10mM NaCl, 3mM MgCl2, 0.1% Igepal CA-630) by being gently pipetted up and down 10 times, and immediately centrifuged for at 1000g for 10 minutes at 4°C.  The tagmentation reaction was performed using the Nextera DNA Library Prep Kit (Illumina-FC-121-1030), with the pellet being resuspended in 50uL 1x TD buffer and with 2.5uL Tn5 Transposase.  This reaction was incubated at 37°C for 30 minutes, after which DNA was purified using MinElute Reaction Cleanup Kit (Qiagen-28204). Tagmented DNA was prepared for Illumina sequencing using the KAPA Real-Time Library amplification kit (KAPA/Roche-KK2701), as per manufacturer's instructions, using Illumina Nextera primers.  Individual samples' reactions were removed upon their qPCR intensity reaching that of Standard 2, which was typically after around 10 cycles of amplification.  Once all samples were sufficiently amplified and removed, the samples were size selected for 150-800bp fragments using AmPureXP beads (Beckman-A63880).  Samples were then sequenced using Illumina's Nextera sequencing primer on the HiSeq 2500 using 100 cycle single read runs.