title: GSM6514323: cDrosha; Caenorhabditis elegans; ncRNA-Seq
library_construction_protocol: Illumina adapter sequences were incorporated to the libraries using PCR. We collected 145 C. elegans pri-miRNAs from MirGeneDB v2.0. We omitted 7 duplicated pri-miRNAs sharing the same miRBase ID. Cel-pri-mir-229 were excluded since its length is longer than 150 nt.   The C. elegans pri-miRNAs were pooled into a substrate mixture.  To clone the cleaved products, the gel pieces containing cleaved products sizing from 50 - 70 nt were sliced out, gel purified, precipitated by IPA, 80% ethanol washed and air dried. The RNAs were dissolved in 10 μL distilled water and ligated with 10 pmol of 3ꞌ-adapter (4N-RA3, /5rApp/NN NNT GGA ATT CTC GGG TGC CAA GG/3ddC/). The ligation was performed in a total 20 μL reaction containing 1 μL  T4 RNA Ligase 2, truncated KQ (NEB, M0373L), 2 μL 10x T4 RNA ligase buffer, 6 μL of 50% PEG8000, and 0.5 μL of SUPERase•In RNase inhibitor (20 U/μL). The mixture was incubated at 25°C for 16 h. After that, it was treated with 20 μL 2x TBE-Urea sample buffer, denatured at 75 °