GSM7850683: ce_smallRNA_lin28_TC_mut_23h; Caenorhabditis elegans; ncRNA-Seq
We harvested worms from 0h to 23h in hourly interval. Worms were collected from liquid culture and washed 3 times and resuspended in Tri Reagent (MRC, TR118) and RNA was extracted using Direct-zol™ RNA MicroPrep kit (Zymo Research; R2062) as per manufacturere's instructions. Total RNA was treated with DNAse on column. There were three sequencing reactions done for this library. First, sequencing libraries were generated using Illumina TruSeq Small RNA Library Prep Kit according to the manufacturer's instructions. The library was sequenced on Illumina HiSeq2500, as 51 cycles single-end reads. since we did not get enough reads, the libraries were reprocessed using CleanTag Small RNA library preparation kit (Trilink Biotechnology) according to manufacturer's instructions. The library pool was size selected on a gel (140-160 bp) to enrich for the small RNA fraction. Subsequently, library was sequenced on NextSeq 500 as 76 cycle single-end reads, and then resequenced on NextSeq 500 as 51 cycle single-end reads as to detect let-7 passenger strand.