adult male thymocytes
Briefly, about 50,000 thymocytes from WT and CKO mice were washed with PBS, and then resuspended in 50 L lysis buffer (10 mM Tris-HCl, pH 7.4, 10 mM NaCl, 3 mM MgCl2, 0.1% Igepal CA-630), and precipitated with centrifugation at 500 g for 10 minutes. The nuclei pellet was washed once with PBS for maximally removing mitochondrial DNA and resuspended in 50 L transposition reaction mixture, which includes 25 L Tagment DNA buffer (Nextera DNA sample preparation kit; Illumina), 2.5L Tagment DNA enzyme (Nextera DNA sample preparation kit; Illumina), and 22.5L l nuclease-free water, and incubated at 37C for 30 minutes. Tagmented DNA was purified with Zymo DNA Clean and Concentrator 5 columns. Based on qPCR amplification results, samples were amplified for additional 4-6 cycles after the 5 cycles pre-amplification (9-11 total cycles) and 1.8 AMPure beads were used for library purification.