GSM7101446: wild-type_rep2; Caenorhabditis elegans; RNA-Seq
Gonad dissections and extractions were performed as in Herbette et al, Cells. 2020 ,doi: 10.3390/cells9092049. Briefly, gonads of 5 to 7 young adults at the L4 stage + 12 h were dissected in dissection buffer (Egg Buffer1.1 × (HEPES pH 7.3 25 mM, NaCl 118 mM, KCL 48 mM, CaCl2 2 mM, 2 mM MgCl2), 0.5 mM Levamisole, 0.1% Tween 20) on slides. Extruded gonads were cut at the elbow and the distal part recovered using a drawn capillary and transferred to 30 µL of ×B extraction buffer (Kit Picopure, Life Technology, # 12204-01, Carlsbad, MA, USA), frozen in liquid nitrogen and stored at −80 °C. For RNA preparation tubes were thawed, the volume of XB extraction buffer adjusted to 100 μL, and RNA purified using the PicoPure kit (Life Technology, # 12204-01) according to the manufacturer's instructions. Elution was in 13 μL of nuclease-free water. The integrity of RNA was evaluated using Tape Station 4200 (Agilent, Santa Clara, CA, USA), and the concentration of RNA measured using DropSense 96 (Unchained Labs, Pleasanton, CA, USA). Librairies were generated using the Stranded Total RNA Prep Ligation with Ribo-Zero Plus kit (Illumina) and sequenced using the Illumina Hiseq 4000 technology (1x50 bases)