title: GSM6761259: ATAC_control_LPS_30_R1; Mus musculus; ATAC-seq
library_construction_protocol: genomic DNA 5 × 104 BMDM cells were pelleted by centrifugation and resuspended in 50 µl of lysis buffer (10 mM Tris-HCl (pH 7.4), 10 mM MgCl2, 0.1% Igepal CA-630), gently pipetting up and down 10 times. Nuclei were pelleted by centrifugation for 10 min at 500g at 4 °C and resuspended in 25 µl of reaction buffer containing 1 µl of Tn5 transposase made in-house using a described protocol (Picelli et al. 2014a), 5 µl of 5x transposase buffer (50 mM Tris-HCl, (pH 8.4) and 25 mM MgCl2), and 19 µl of milliQ water. The tagmentation mix was incubated at 37 °C for 60 min gently shaking, in order to avoid precipitation of nuclei. The reaction was stopped by the addition of 5 µl of cleanup buffer (900 mM NaCl, 30 mM EDTA), 2 µl of 5% SDS and 2 µl of Proteinase K (20 µg /µl) (Merck cod. P2308) and incubation for 30 min at 40 °C. Tagmented DNA was isolated using 2× Agencourt Ampure Xp beads (Beckman cod.A63881) and amplified by PCR with the HotStart Kapa HiFi enzyme (Roche cod. 07958889001). Fragments smaller than 600 bp were isolated by negative size selection (using 0.65× Agencourt Ampure Xp beads) and then purified with 1.8× Agencourt Ampure Xp beads. Libraries were single-end sequenced (76 bp) on an Illumina NextSeq500 platform.