GSM8591250: SS-NAM-3; Mus musculus; OTHER
Flow-sorted nerve- and airway-associated macrophages A total of 50,000 purified macrophage populations per replicate were washed once in cold sterile PBS and resuspended in 20-50 µl of cold lysis buffer (10 mM Tris-HCl (pH 7.4), 10 mM NaCl, 3 mM MgCl2 and 0.1% IGEPAL CA-630). Cell lysates were centrifuged at 500g for 30 seconds to 1 min at 4 °C, and nuclei were resuspended in 25 µl of water. The transposition reaction mix consisted of 25 µl of TD buffer, 2.5 µl of Tn5 transposase (Illumina cat. 20034198), and 22.5 µl of nuclease-free water containing the nuclei. The mix was incubated for 30 min at 37 °C. Transposed DNA fragments were purified using a Qiagen Reaction MiniElute kit (Qiagen). Clean DNA was run with PCR amplification to incorporate the indexes, which were a universal index and a unique i7 index per sample. (IDT primers outlined in Buenrostro 2013 paper) and amplified by PCR for 11 cycles using NEBNext high fidelity 2× PCR master mix (New England Biolabs).  PCR products were cleaned with 0.9x volume of Ampure XP beads (Beckman Coulter), and amplified fragment size was verified on an Agilent 2100 Bioanalyzer system (Agilent) with high sensitivity DNA screentape. ATAC-seq libraries were quantified by quantitative PCR with the Roche-kapa Illumina quantification kit. Paired-end sequencing was performed on an Illumina Novaseq6000 system with an SP100 cycle kit sequenced as paired-end 50 bases.