Researchers at the University Leipzig and the University of Vienna have developed a dedicated mapping strategy to handle RNA-seq reads that map to tRNAs relying on a modified target genome in which known tRNA loci are masked and instead intronless tRNA precursor sequences are appended as artificial “chromosomes”. In a first pass, reads that overlap the boundaries of mature tRNAs are extracted. In the second pass, the remaining reads are mapped to a tRNA-masked target that is augmented by representative mature tRNA sequences. Using both simulated and real life data the researchers show that their best-practice workflow removes most of the mapping artefacts introduced by simpler mapping schemes and makes it possible to reliably identify many of chemical tRNA modifications in generic small RNA-seq data. Using simulated data the FDR is only 2%. They find compelling evidence for tissue specific differences of tRNA modification patterns.
Scheme of the best-practice workflow for detecting tRNA misincorporations
The top part describes the construction of the artificial and cluster genome, the middle section refers to the mapping steps, the bottom layer describes the post-processing.
Availability – The workflow is available both as a bash script and as a Galaxy workflow from https://github.com/AnneHoffmann/tRNA-read-mapping.