title: Small RNA-seq of Caenorhabditis elegans young adults: tofu-2 point mutant, pid-1 mutant, experiment 2, replicate 3
design: L1 worms were seeded on NGM OP50 plates and grown at 20 C until young adult stage, washed with M9 buffer and snap frozen in liquid nitrogen in 50 ul aliquots. For RNA purification, worm aliquots were thawed on ice, 500 ul of TRIzol LS Reagent (Termo Fisher) was added and samples underwent five freezing-thawing cycles in liquid nitrogen followed by centrifugation. Supernatants were collected and RNA purification was carried out with Direct-zol RNA Microprep Kit (Zymo Research) according to its protocol. NGS library prep was performed with NEXTflex Small RNA-Seq Kit V3 following Step A to Step G of Bioo Scientific`s standard protocol (V19.01). Libraries were prepared with a starting amount of 340 ng and amplified in 16 PCR cycles. Amplified libraries were purified by running an 8% TBE gel and size-selected for 15-40 bp insert length. Libraries were profiled in a High Sensitivity DNA Chip on a 2100 Bioanalyzer (Agilent Technologies), quantified using the Qubit dsDNA HS Assay Kit, in a Qubit 2.0 Fluorometer (Life Technologies), and sequenced on Illumina NextSeq 500 sequencer.
library_name: TOFU2_PID1_exp2_rep3