title: GSM6045696: ATAC-seq in Lola-I mutant ORC4 Trachea 14-17hrs embryo Rep2; Drosophila melanogaster; ATAC-seq
library_construction_protocol: Nuclei were isolated from INTACT embryos 14-17h AED by resuspending and douncing embryos in HBS buffer (0.125 M Sucrose, 15 mM Tris (pH 7.5), 15 mM NaCl, 40 mM KCl, 2 mM EDTA, 0.5 mM EGTA). The nuclei were further dissociated using syringe (22.5 gauge needle) 10 times. After washins with HBS buffer, nuclei were incubated with Dynabeads® M-280 Streptavidin beads( Invitrogen, # 11205D) for 30 minutes with end to end rotation at 4°C. A magnet was used to separate the bead bound nuclei, and the beads were washed thoroughly with HBS buffer. Purity of nuclei isolation was determined by staining nuclei with DAPI, and presence of free nuclei, unbound to beads, were used as indicators of contamination. Isolated nuclei were treated with Tn5 transposase((Illumina, #FC-121-1030). Nextera index kit (Illumina, #FC-121-1011) was used to create libraries.