ATAC-seq for Chromatin Accessibility

For the global chromatin accessibility experiments ductal and lobular cell lines were cultured for three days in HD conditions and then treated with 10nM estradiol for 45 minutes. Nuclei of 100,000 freshly fixed in 1% formaldehyde cells were processed as previously reported (15 (link)). Briefly, cells were resuspended in 1mL of cold ATAC-seq resuspension buffer (RSB; 10mM Tris-HCl pH 7.4, 10mM NaCl, and 3mM MgCl₂ in water) and centrifuged. Cell pellets were then resuspended in ATAC-seq RSB (0.1% NP40, 0.1% Tween-20, and 0.01% digitonin) and incubated on ice. After lysis, ATAC-seq RSB containing 0.1% Tween-20 (without NP40 or digitonin) was added. Nuclei were centrifuged and then were resuspended in 50μl of transposition mix (25μl 2× TD buffer, 2.5μl transposase (100nM final), 16.5μl PBS, 0.5μl 1% digitonin, 0.5μl 10% Tween-20, and 5μl water). Transposition reactions were incubated at 37°C for 30 min. Reactions were cleaned up with Zymo DNA Clean and Concentrator5 columns.