ATAC-seq from Duodenal Villi

Duodenal villi were treated with pre-warmed 0.25% Trypsin for 8 min at 37°C on a vortex station (speed set between 6–7), neutralized with 10% FBS, and passed through a 40-μm cell strainer to obtain single cells. 50,000 cells were used for ATAC-seq as described previously^{37 ,38 (link)} with slight modifications. Briefly, cells were centrifuged at 500 g for 5 min at 4°C and resuspended in ice-cold lysis buffer (10 mM Tris, pH 7.4, 10 mM NaCl, 3 mM MgCl₂, and 0.1% NP-40). Cells were then centrifuged at 500 g for 10 min at 4°C. The isolated nuclear pellets were incubated with a 50-μl reaction of Nextera Tn5 Transposase (Illumina FC-121–1030) for 30 min at 37°C. The transposed chromatin was purified with QIAquick PCR Purification Kit (QIAGEN), and PCR was amplified with high-fidelity 2× PCR Master Mix (New England Biolabs M0541). The optimum number of additional cycles was based on 1/3 of the maximum fluorescent intensity. The PCR amplified libraries were purified. Fragment size was selected using Pippin Prep and sequenced on Illumina NextSeq 550.