title: GSM4774870: ISC young rep3 [ATAC-Seq]; Drosophila melanogaster; ATAC-seq
library_construction_protocol: For ATAC-seq analysis, the protocol described in (Buenrostro, Wu, Chang, & Greenleaf, 2015) was used with minor adjustments. For each ATAC-seq sample, 100,000 YFP+ ISCs were sorted by FACS and cells were immediately spun down at 4  ̊C, washed, and then lysed and released nuclei were used in a transposition reaction using Nextera DNA Sample prep kit according to manufacturer's protocol (Illumina). Transposed DNA was purified using MiniElute Reaction Cleanup kit (Qiagen) and eluted in 10 μl water. Purified transposed samples were amplified using NEBNext High-Fidelity 2X PCR Master Mix (New England Biolabs) per manufacturer's user guide. Generated libraries were quantified by qPCR to determine the cycle number at which each sample produced 25% of maximum fluorescent intensity. The fluorescent intensity was used to estimate the number of additional PCR cycles required for each sample. Libraries were purified using SPRIselect beads (Beckman Coulter) to remove contaminating primer dimers. Library quality was assessed with dsDNA HS Assay kit (Thermo Fisher) and quantitated by Qubit 3.0 Fluorometer (Thermo Fisher). Pooled libraries were quantitated by Kapa Library Quantification Kit (Kapa Biosystems) and sequenced on Illumina HiSeq2500 to generate 30 M of pair end 50 bp reads per library.