GSM5825251: ZT6_18C_WT_Rep2; Drosophila melanogaster; RNA-Seq
To collect dissected fat body samples at 2-hour and 4-hour intervals from ZT0 (set to 8AM) to ZT22 set (set to 10PM) during the daytime only, flies were entrained in two separate incubators with inverse light:dark schedule (LD and DL). On the day of sampling, flies were directly dissected without dry ice to harvest fat tissues in the abdomen. Pinned flies were cut to remove organs in the abdomen (intestine, ovaries, malpighian tubules, etc). Fat tissues attached to epidermis was collected. Dissected fat body tissues from 10 flies were homogenized in pH 7.4 PBS for 2 min using a Kontes motor and pestle, and were incubated TRIzol LS Reagent (Thermo Fisher, Waltham, MA) for 15 min. RNA was extracted according to the manufacturer's instructions and residual DNA in the homogenized samples was removed by RNAse free DNase I (Thermo Fisher Scientific). Quality of RNA samples were checked with Agilent 2100 Bioanalyzer. cDNA library was constructed with poly(A) selected mRNA using Truseq RNA library preparation kit and then sequenced at the Genomics Core Facility at the University of Chicago on Illumina HiSeq 2000 System.