GSM8970410: RNA extracted from C. elegans grown without ethanol was analyzed at time zero (biol rep 4).; Caenorhabditis elegans; RNA-Seq
Total RNA was extracted from whole C. elegans L3 larvae using TRI Reagent® (Molecular Research Center, Inc., TR118) according to the manufacturer's instructions. Approximately 1,500 synchronized L3 larvae were collected after ethanol treatment and homogenized in 0.5 mL TRI Reagent® using a plastic pestle. After homogenization, 50 µL of 1-bromo-3-chloropropane was added, followed by vigorous shaking for 15 seconds. Samples were incubated at room temperature for 15 minutes and centrifuged at 12,000 × g for 15 minutes at 4°C. The aqueous phase was transferred to a new tube and RNA was precipitated with isopropanol, followed by washing with 75% ethanol. The RNA pellet was air-dried and dissolved in RNase-free water. Libraries were prepared with Kapa RNA mRNA HyperPrep kit (Roche) according to the manufacturer's protocol.