GSM8640579: nspc_tent-5_2; Caenorhabditis elegans; RNA-Seq
Populations of worms were collected and washed three times with 50 mM NaCl. Worm pellets were resuspended in 1 ml TRI Reagent (Sigma-Aldrich), vortexed for 15 minutes at room temperature, and stored at -80°C until further processing. RNA extraction was performed according to the manufacturer's protocol for TRIzol-based isolation, with additional purification using KAPA Pure magnetic beads (RNA to beads ratio: 1:3 v/v). RNA quality and integrity were confirmed using the Agilent TapeStation system. Total RNA samples for Illumina RNA sequencing were sent to Center for New Technology (Poland) or Genewiz (Germany) and libraries were prepared with the use of internal protocols. Sequencing libraries for Nanopore direct RNA sequencing were prepared using the DRS Kit (Oxford Nanopore Technologies, SQK- RNA002) according to the manufacturer's protocol. In each library, total RNA isolated from worms (1,5 - 2 μg depending on the amount of isolated RNA) was mixed with 2 ng of in vitro transcribed poly(A) standards.