GSM8443986: MEFs treated with doxycycline, Day 2, ATAC-seq; Mus musculus; ATAC-seq
ATAC-seq libraries from different timepoints were prepared starting from 50,000 cells, following previously published protocols (Buenrostro et al, 2013, 2015; Ackermann et al, 2016; Corces et al, 2017). Briefly, cell pellets were lysed in Lysis Buffer [10mM Tris-HCl, pH 7.5, 10mM NaCl, 3mM MgCl2, 0.1% NP-40, 0.1% Tween-20, 0.01% Digitonin (Promega, G9441)] for 3 min on ice, washed with Wash Buffer [10mM Tris-HCl, pH 7.5, 10mM NaCl, 3mM MgCl2, 0.1% Tween-20] and centrifuged at 500g for 10 min at 4o C. Each pellet (nuclei) was resuspended in 50ul Transposition reaction mix [1x Tagment DNA Buffer (Illumina, 15027866), 16.5ul 1x PBS, 0.1% Tween-20, 0.01% Digitonin (Promega, G9441), 2.5ul Tn5 Transposase (Tagment DNA Enzyme 1, Illumina, 15027865)] and incubated at 37o C for 30 min on a thermomixer at 1000rpm. DNA purification was performed using the Qiagen MinElute Reaction Cleanup Kit (Qiagen, 28204). Library purification was performed with the AMPure XP beads (Beckman Coulter, A63880). The quality of libraries was assessed using an Agilent 2100 Bioanalyzer and libraries were quantified with the Qubit™ High Sensitivity (HS) spectrophotometric method (Invitrogen). Libraries were sequenced paired-end using a NovaSeq 6000 sequencer at the Greek Genome Center at BRFAA.