GSM5966503: Sample 12: Post-PPx: Arid1aFl/Fl; MIP-rtTA control; Mus musculus; ATAC-seq
40,000 isolated islet cells in single cell suspension were used to prepare crude nuclei, which were immediately used for the transposition reaction following a published protocol (Buenrostro et al. 2015). Transposition reaction using Nextera Tn5 transposase (Nextera Tn5 Transposase, Illumina #FC-121-1030) was incubated at 37 °C for 30 min with gentle shaking. DNA was eluted using Qiagen MinElute PCR Purification kit and transposed DNA fragments were amplified using Nextera PCR Primer 1 and Primer 2 (containing barcode). Thermal cycle settings were as follows: 1 cycle of 72°C for 5 min, 98°C for 30 sec, 5 cycles of 98°C for 10 sec, 63°C for 30 sec, 72°C for 1 min. 5-6 additional cycles were used as determined for each sample with a qPCR reaction to avoid artifacts due to over-amplification. Amplified library was purified using Qiagen MinElute PCR Purification kit. Amplified libraries were size selected using Ampure XP beads and quality was analyzed using Bioanalyzer High Sensitivity DNA kit. Indexed libraries were multiplexed in a single flow cell and underwent 75 base pair single-end sequencing on an Illumina NextSeq500 using the High Output kit v2 (75 cycles) 75 base pair single-end sequencing on an Illumina NextSeq500 using the High Output kit v2 (75 cycles)