GSM4910666: Input-RELACS.dom-kd.st5_rep2; Drosophila melanogaster; OTHER
RELACS protocol was performed as described previously by Arrigoni, et al. 2018. Briefly, embryos were thawed in RELACS lysis buffer (10 mM Tris-HCl [pH 8], 10 mM NaCl, 0.2% Igepal, 1x Protease inhibitor cocktail) and the nuclei were isolated by sonication using the NEXSON procedure(Arrigoni et al., 2016). To digest the chromatin, 25 µl of 10x CutSmart buffer (NEB), 2.5 µl 100x Protease inhibitor cocktail and 1 µl of CviKI-1 (50 U/µl, NEB R0710S) were added. The digestion reaction was incubated overnight at 20 °C. End repair and A-tailing was performed and customized adapters (Arrigoni et al., 2018) were ligated to the fragments. Once barcoded, the samples were pooled together. Chromatin was then sheared by sonication (Covaris E220, MicroTubes, 5min, peak power 105, duty factor 2, cycles burst 200). This chromatin was used for automated ChIP with the IP-Star Diagenode system. IPs and Inputs were decrosslinked and DNA was purified. Libraries were prepared using the NEB Ultra II DNA Library Prep Kit for Illumina (E7645S and E6440) following the manufacturer's instructions