GSM4563060: T2DMEF2; Mus musculus; ATAC-seq
Post-FACS cells were applied directly to the Fast-ATAC optimized ATAC-seq workflow (Corces et al. 2016), with the following modifications. Cells were washed 1x in MEF media, and then 75,000 cells were resuspended in 1 mL warm RPMI media (Gibco). Following a 5 min spin at 4˚C (500 x g), cells were washed with 1 mL ATAC-RSB (10 mM Tris-HCl pH 7.4, 10 mM NaCl, and 3 mM MgCl2 + 0.1% Tween), and then resuspended in TDE1 transposase solution (Illumina) + 0.01% digitonin. The transposase mix was incubated at 37˚C with shaking for 30 mins, after which the transposed DNA was purified using a MinElute Reaction Cleanup Kit (QIAGEN). Transposed fragments were amplified for 8 cycles using indexed primers according to Buenostro et al 2015, and then purified by AMPure XP bead selection (Beckman Coulter). ATAC libraries were quantified using the NEBNext Library Quant Kit (NEB) and assayed using a Bioanalyzer High Sensitivity DNA chip (Agilent) to establish library quality. Pooled libraries were sequenced on an Illumina NextSeq 500 using a High Output Kit v2.5 (75 cycles) (Illumina).