GSM7415392: vasGFP_strain_dmel_6_ovaries (ATAC-seq); Drosophila melanogaster; ATAC-seq
For the cell lines, the omni-ATAC-seq protocol was performed using 50,000 cells as described before. For the ovaries, the freshly dissected instact ovaries were directly lysed in ATAC-Resuspension Buffer (RSB) containing 0.1% NP40, 0.1% Tween20, and 0.01% Digitonin and washed out with cold ATAC-RSB containing 0.1% Tween-20. The transposition reaction was performed with 25 μl 2x TD buffer from and 2.5 μl transposase (100nM final) from Illumina Tagment DNA Enzyme and Buffer Small Kit (Illumina 20034197), 16.5 μl PBS, 0.5 μl 1% digitonin, 0.5 μl 10% Tween-20, 5 μl H2O at 37°C for 30 minutes in a thermomixer with 1000 RPM mixing. Zymo DNA Clean and Concentrator-5 Kit (cat# D4014) was used for a clean-up. The transposed fragments were amplified for 5 cycles [72 °C for 5 min, 98 °C for 30 s (98 °C for 10 s, 63 °C for 30 s, 72 °C for 1 min) × 5] using NEBNext 2x MasterMix and primer pairs of the universal primer Ad1 and the index primers Ad2. This was followed by qPCR amplification to determine additional cycle number. Amplified DNA was purified using Zymo DNA Clean and Concentrator-5 Kit (cat# D4014) and eluted in 20 μl H2O. AMPure XP beads (Beckman Coulter) were used for double-sided bead purification. 100-600 bp fragments were selected using Blue Pippin (Sage Science).