GSM3290162: ATAC-seq_LV4 (P13); Mus musculus; ATAC-seq
Next, the cell pellets were resuspended in 50 µl of ATAC-lysis buffer (10mM Tris HCl pH7.4, 10mM NaCl, 3mM MgCl2, 0.1% Tween-20, 0.1% NP40, and 0.01% Digitonin) and incubated on ice for 3 min. Wash out lysis with 1 ml of cold ATAC-lysis buffer containing 0.1% Tween-20 but No NP40 or digitonin and invert tube 3 times to mix. Nuclei were pelleted at 500 RCF for 10 min at 4°C in a fixed angle centrifuge. Cells nuclei were resuspended in 50 µl of transposition mixture (25 ul 2x TD buffer, 2.5 ul transposase (100 nM final), 16.5 ul PBS, 0.5 ul 1% digitonin, 0.5 ul 10% Tween-20, and 5 ul H2O) (Nextera DNA Sample Preparation Kit, Illumina, FC-121-1030). The reaction was performed  at 37°C for 30 minutes in a thermomixer with 1000 RPM mixing. The transposed DNA was purified using a Zymo DNA Clean and Concentrator-5 Kit (D4014). DNA libraries were PCR amplified using NEBNext High-Fidelity 2x PCR Master Mix (Bioke, M0541), and size selected for 200 to 800 bp using homemade Serapure beads [60]. Library concentrations were quantified with the KAPA Library Quantification Kit (KK4854), and equimolar amounts were pooled for single-end sequencing on an Illumina HiSeq 4000 instrument (Illumina) to yield ~50 million (range 34-90 million) 51bp long reads per sample.