GSM2070433: lin-65(n3441) EV replicate 3; Caenorhabditis elegans; RNA-Seq
Collected worms were snap frozen in liquid nitrogen. Pellets were grinded with a mortar and pestle on dry ice and the resulting powder was thawed in the presence of lysis buffer (50mM Tris-HCl, 50mM NaCl, 10mM MgCl2, 5mM CaCl2 200μg/ml cycloheximide, 200μg/ml heparin, 1% Triton X-100, 0.1% NaDOC) and incubated on ice for 10 minutes prior to centrifugation at 4C, 16,000 x g for 10 minutes.  Total RNA was purified from clarified lysate using Trizol-LS (Life Technologies) and poly-A RNA was extracted with oligo-dT(25) Dynabeads (Ambion), each according to manufacturer’s instructions. Sequencing libraries were prepared using the Epicentre ScriptSeq v2 according to manufacturer’s instructions.