GSM3674197: Day5_sgFOXA1_2_rep1; Mus musculus; ATAC-seq
At time of collection, cells were trypsinzed, and 50,000 cells (counted by using trypan blue exclusion) were processed for ATAC-sequencing as follows.  After a wash step in cold Cell Wash Buffer (CWB= 10 mM Tris-HCl pH 7.4, 10 mM NaCl, 3 mM MgCl2), outer membranes were disrupted in lysis buffer (CWB + 0.1% NP40) for 2min on ice. Lysis reaction was stopped with the addition of 1ml of CWB. After a centrifugation step 1,500g for 10min, pelleted nuclei are kept for the next step. In a 50μl final volume, tagmentation was performed for 30min at 37C, using the kit Nextera DNA library prep kit (Illumina cat# FC-121-1030). After addition of SDS 0.2% final concentration, DNA is purified in AMPure XP beads (Beckman Coulter cat# A63881) using a ratio 2:1 (V/V) beads:tagmented DNA.Freshly eluted DNA was barcoded and amplified in 110μl PCR volume (NEB Next Q5 Hot Start HiFi PCR, cat# M0543L) to generate library with the following PCR program: 65C, 5min, 98C, 30sec, (98C, 10sec – 65C, 30sec) *11cycles, 4C hold.
GEO Accession
GSM3674197