Ovary and testis were dissected from age-matched adult flies of the genotypes w1118; tra2ts2/tra2ts1 (experimental) or w1118 (control for dsxF->dsxM experiments); for dsxM->dsxF experiments the genotypes were: y1 w*; P{w+mc=UAS-Tra.F}20J7; P{w+mc=tubP-GAL80ts}7/P{w+mc=tubP-GAL4}LL7 (experimental) or P{w+mc=tubP-GAL80ts}7/P{w+mc=tubP-GAL4}LL7 (control). All samples were raised at 18°C until 5 days after eclosion when adults were shifted to either 29°C (for tra2ts) or 30°C (for UAS-TraF) for 0, 12, or 24 hours.  Total RNA was extracted from ovary and testis dissected at room temperature (placed on ice after 30 minutes) using TRIzol Reagent following manufacturer’s protocol (Ambion Life Technologies, Carlsbad, CA, USA).  Egg chambers stage 10 and older were removed and discarded to enrich for younger-staged egg chambers. Purified RNA was treated with DNase I following manufacturer's protocol (New England Biolabs, Ipswich, MA, USA) and purified again using phenol:chloroform extraction followed by ethanol precipitation.  Duplicate RNA-seq libraries were constructed from 200 ng total RNA from independent dissection of each sample using the TruSeq RNA Sample Preparation v2 high-throughput (HT) protocol (Illumina, San Diego, CA, USA, 2011).  Libraries were sequenced on the HiSeq 2000 machine following a 76 bp single-end protocol (Illumina, San Diego, CA, USA). True-Seq v2 protocol starting with 200 ng of total RNA.
GEO Accession
GSM2091025