title: Caenorhabditis elegans wild-type strain N2 Bristol, larval stage L4, direct RNA sequencing, biological replicate 1
library_construction_protocol: The synchronized wild-type C. elegans strain (N2 Bristol) were kept at 20°C. Worms were collected from plates 10 h after entering the L4 stage and washed three times using 50 mM NaCl. Worm pellet was then suspended in 1 ml of TRIReagent (Sigma-Aldrich; cat. no: T9424), vortexed for 15 min in room temperature and stored in -80°C before the RNA isolation. RNA was isolated following the manufacturer's instructions and cap-enrichment of mRNA was performed as described in Bilska et al. (2020). Nanopore direct RNA sequencing libraries were prepared with a Direct RNA Sequencing Kit (SQK-RNA002; ONT) from 4 µg of cap-enriched mRNA. 150 ng of oligo-(dT)25-enriched mRNA from Saccharomyces cerevisiae and 2 ng of standards with predefined poly(A) lengths were added to each library as described in Bilska et al. (2020).