Figure 1. Experimental and bioinformatic pipeline for tRNA annotation and reference transcript curation by hydro-tRNAseq.

(A) tRNAs and pre-tRNAs were size-selected from HEK293 total RNA and subjected to limited alkaline hydrolysis, followed by dephosphorylation, rephosphorylation and conventional small RNA sequencing as described previously (Hafner et al., 2012b). (B) An iterative mapping and annotation protocol was used to first annotate and curate fully processed and nucleotide-modified mature tRNAs. Leftover reads that spanned the mature-precursor junctions were used to identify transcribed tRNA genes. See also Fig. S1, S2, S8 and Tables S1 and S2.