library_construction_protocol: Cells were harvested and shock frozen, extraction was performed by Active Motif (1. Prepare nuclei. To prepare nuclei, we spun 50,000 cells at 500g for 5 min, which was followed by a wash using 50 μL of cold 1× PBS and centrifugation at 500g for 5 min. Cells were lysed using cold lysis buffer (10 mM Tris-HCl, pH 7.4, 10 mM NaCl, 3 mM MgCl2 and 0.1% IGEPAL CA-630). Immediately after lysis, nuclei were spun at 500g for 10 min using a refrigerated centrifuge. To avoid losing cells during the nuclei prep, we used a fixed-angle centrifuge and carefully pipetted away from the pellet after centrifugations. 2. Transpose and purify. Immediately following the nuclei prep, the pellet was resuspended in the transposase reaction mix (25 μL 2× TD buffer, 2.5 μL transposase (Illumina) and 22.5 μL nuclease-free water). The transposition reaction was carried out for 30 min at 37 °C. Directly following transposition the sample was purified using a Qiagen MinElute kit). Following purification, we amplified library fragments using 1× NEBnext PCR master mix and 1.25 μM of custom Nextera PCR primers 1 and 2 (Supplementary Table 1), using the following PCR conditions: 72 °C for 5 min; 98 °C for 30 s; and thermocycling at 98 °C for 10 s, 63 °C for 30 s and 72 °C for 1 min. To reduce GC and size bias in our PCR, we monitored the PCR reaction using qPCR in order to stop amplification before saturation. To do this, we amplified the full libraries for five cycles, after which we took an aliquot of the PCR reaction and added 10 μl of the PCR cocktail with Sybr Green at a final concentration of 0.6×. We ran this reaction for 20 cycles to determine the additional number of cycles needed for the remaining 45-μL reaction. The libraries were purified using a Qiagen PCR cleanup kit yielding a final library concentration of ∼30 nM in 20 μL. Libraries were amplified for a total of 10–12 cycles. During the ATAC-seq protocol, we chose to avoid a size-selection step to maximize the library complexity. The sequenced insert size is a distribution between 40 bp and 1 kb with a mean of ∼120 bp. From the bioanalyzer and gels we observed fragments >2 kb, which would make Qubit and other mass-based quantitation methods hard to interpret. For this reason we quantified our libraries using qPCR-based methods.