title: D_sechellia_females
design: To collect RNA sequencing data, D. simulans (wXD1), D. mauritiana (w12) and D. sechellia  (Rob3c / Tucson 14021-0248.25) were reared at room temperature in vials on standard cornmeal media or standard cornmeal-molasses medium. Males and virgin females were collected post-eclosion and allowed to age for 3-5 days in single-sex vials.  20-30 flies were then flash frozen in liquid nitrogen and stored at -80C for RNA extraction. For D. sechellia, total RNA was isolated from whole tissue using the RNeasy Plus Kit (Qiagen, Valencia, CA) per manufacturers recommendations. The TruSeq RNA Sample Preparation Kit V2 (Illumina, San Diego, CA) was used for next generation sequencing library construction per manufacturers protocols. Briefly, polyA mRNA was purified from ~100 ng total RNA with oligo-dT magnetic beads and fragmented. First-strand cDNA synthesis was performed with random hexamer priming followed by second-strand cDNA synthesis. End repair and 3' adenylation was performed on the double stranded cDNA. Illumina indexed adaptors were ligated to both ends of the cDNA, purified by gel electrophoresis and amplified with PCR primers specific to the adaptor sequences to generate amplicons of approximately 200 to 500 bp in size. The amplified libraries were purified by AMPure (Beckman Coulter, Brea, CA) purification and hybridized to an Illumina pair end flow cell for cluster amplification using the cBot (Illumina, San Diego, CA) at a concentration of 8 picomoles per lane.