GSM6260856: HSBAM1T_S7_R1_001; Drosophila melanogaster; OTHER
The ovaries were immediately flash frozen on liquid nitrogen. Samples were homogenized by motorized pestle in lysis buffer and 20% of lysate was used as input for mRNA isolation and library preparation (as described above). Samples were loaded onto 10-45% CHX supplemented sucrose gradients in 9/16 x 3.5 PA tubes (Beckman Coulter, #331372) and spun at 35,000 x g in an SW41 rotor for 3 hours at 4°C. Gradients were fractionated with a Density Gradient Fractionation System (#621140007). RNA was isolated using TRIzol (Invitrogen, 15596026) or phenol-chloroform treatment, treated with DNase (TURBO DNA-free Kit, Life Technologies, AM1907).  mRNA-enriched libraries were obtained by treating total RNA with poly(A)tail selection beads and then following the manufacturer's instructions of the NEXTflex Rapid Directional RNAseq Kit (Bioo Scientific Corp, NOVA-5138-08).