GSM7835199: tRNA-NT1; Homo sapiens; ncRNA-Seq
For tRNA-seq, total RNA was extracted using TRIzol Reagent (#15596018; Life Technologies) and chloroform/isopropanol precipitation method. Next, tRNA was isolated from total RNA using denaturing urea polyacrylamide gel electrophoresis (Urea PAGE) and then subjected to m1A & m3C demethylation using rtStarTM tRF&tiRNA Pretreatment Kit (#AS-FS-005; Arraystar). Demethylated tRNA was purified and then partially hydrolysed according to Hydro-tRNAseq method. Small RNA sequencing libraries were constructed using NEBNext® Multiplex Small RNA Library Prep Set for Illumina® kit (E7300L/E7850L; New England Biolabs) by Arraystar. Size selection of ~140-155 bp PCR amplified fragments (corresponding to ~19-35 nt tRNA fragments size range) was performed. The produced tRNA-seq libraries were qualified and absolutely quantified and sequenced on Illumina sequencer.