RNA extraction was performed as previously outlined (Mossman, Tross et al. 2016 (link)). Briefly, we extracted RNA from 30, 5-day-old healthy flies in each genotype x hypoxia × sex treatment. There were three replicates per genotype × hypoxia × sex condition, with the exception of one sample of siI;AutW132 females, which had two replicates at hypoxia timepoint = 0; resulting in a total of 71 RNA libraries. RNA extraction followed a modified RNA-Seq sample preparation protocol from the Gilad Laboratory (Marioni et al. 2008 (link)). Messenger RNA was first extracted, followed by RNA fragmentation, cDNA first strand synthesis, second strand synthesis, end repair, poly adenylation, adapter ligation and PCR enrichment. Throughout, RNA and DNA were quantified using the Qubit® Kits (RNA Broad Range, ds DNA Broad Range, and ds DNA High Sensitivity) with a Qubit® 1.0 Fluorometer. All Qubit® reagents were obtained from Molecular Probes™ (ThermoFisher Scientific). Following PCR enrichment, we size selected PCR products with size range = 334–500 bp using a Caliper Lab Chip XT (DNA 750 chip) (Caliper Life Sciences, Inc. Hopkington, MA, USA).