!Citation=Wittes et al. 2018. [submitting] !Title=A gene expression screen in Drosophila melanogaster identifies novel JAK/STAT and EGFR targets during oogenesis !PubMedID= !Name=Drosophila egg chambers: control vs. ectopic JAK/STAT and EGFR signaling !ExptSetNo=7324 !Description=Summary: Gene expression analysis was performed to identify transcripts enriched in egg chambers with ectopic JAK/STAT and EGFR signaling in the follicle cells. JAK/STAT and EGFR were ectopically activated by crossing GAL80TS ; GR1-GAL4 flies to UAS-Unpaired, UAS-lambdatorpedo flies to obtain the genotype GAL80TS/+ ; GR1-GAL4/UAS-Unpaired, UAS-lambdatorpedo. As a control, GAL80TS ; GR1-GAL4 flies were crossed to OreR (WT) flies to obtain flies of genotype GAL80TS/+ ; GR1-GAL4/+. Ectopic JAK/STAT and EGFR activation generates extra posterior follicle cels (PFCs), and the goal of this experiment was to identify potential PFC-enriched transcripts. Growth protocol: Crosses were carried out at 18 degrees Celsius. Adult females of specified genotypes were fed yeast for two days prior to dissection. Transgene expression was induced by incubating flies at 29 degrees Celsius for 16 hours before dissecting out whole ovaries. Overall design: A reference design was used to control for dye bias. Reference (WT) RNA was isolated from the ovaries of OreR flies. Biological replicates: 3 control replicates, 3 experimental replicates. Extraction protocol: RNA was extracted from ovaries using TRIzol reagent and further purified with Qiagen RNeasy columns. It was then treated with TURBO DNase. Label protocol: Each labeling reaction was performed with 322 ng of RNA. Universal reference RNA was labeled with Cy3-CTP and all other RNA samples were labeled with Cy5-CTP. Labeling was performed with the Quick Amp two-color labeling kit. Labeled cRNA was purified by RNeasy column. Dye incorporation and cRNA yield were determined by NanoDrop. Hybridization protocol: Labeled cRNA was hybridized for 17 hours at 65 degrees Celsius. Hybridization and washes were performed according to Quick Amp kit protocols. Scanning protocol: Slides were scanned on an Agilent G2505C scanner at a 5 micron resolution. Images were quantified using Agilent Feature Extraction Software (version A.8.5.1.1).