GSM2127816: Anterior_ovariole_Day3_Replicate2; Drosophila melanogaster; RNA-Seq
Flies were collected and aged as described above and ovaries were dissected in a glass embryo dish in ice cold Phosphate Buffer Saline (PBS). Ovaries were immediately transferred to fresh ice cold PBS in a new dish. The ovarioles were teased apart using tungsten needles. Germaria and round previtellogenic egg chambers up to stage 8 were   separated from the rest of the ovary using tungsten needles and were immediately transferred to RNAlater (Life Technologies, Carlsbad, CA) on ice. Dissected samples were frozen at -80°C until RNA extraction, and each sample had 30 to 50 ovarioles. Three biological replicates were prepared for each age group. RNA was prepared after all the samples were dissected. Total RNA was isolated using the RNeasy mini kit (Qiagen, Valencia, CA) following manufacturer’s protocol with the following modification. RNAlater solution was removed from the tissue and the tissue was placed in 600 μl Buffer RLT (supplied with the kit). The tissue was then disrupted using five to ten glass beads in a mini-beadbeater (Biospec Products, Bartlesville, OK) for 2 rounds of 30 sec each. After this point we followed the manufacturers protocol exactly. RNA was quantified using Quant-iT Ribogreen RNA assay kit (Invitrogen, Carlsbad, CA) following manufacturers protocol and 200 ng of RNA was used for library preparation. Libraries were prepared according to Lin et al. (BMC Genomics, 2016, PMID: 26732976) except we used 200 ng total RNA per sample as input.
GEO Accession
GSM2127816