mim-tRNAseq overcomes experimental and computational hurdles to tRNA quantitation
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mim-tRNAseq includes a comprehensive computational toolkit for tRNA read analysis
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tRNA abundance, aminoacylation, and modification status quantified in one reaction
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mim-tRNAseq reveals an interdependence of modifications at distinct tRNA positions
Summary
Measurements of cellular tRNA abundance are hampered by pervasive blocks to cDNA synthesis at modified nucleosides and the extensive similarity among tRNA genes. We overcome these limitations with modification-induced misincorporation tRNA sequencing (mim-tRNAseq), which combines a workflow for full-length cDNA library construction from endogenously modified tRNA with a comprehensive and user-friendly computational analysis toolkit. Our method accurately captures tRNA abundance and modification status in yeast, fly, and human cells and is applicable to any organism with a known genome. We applied mim-tRNAseq to discover a dramatic heterogeneity of tRNA isodecoder pools among diverse human cell lines and a surprising interdependence of modifications at distinct sites within the same tRNA transcript.