Many aspects of the RNA maturation leave traces in RNA sequencing data in the form of deviations from the reference genomic DNA. This includes, in particular, genomically non-encoded nucleotides and chemical modifications. The latter leave their signatures in the form of mismatches and conspicuous patterns of sequencing reads. Modified mapping procedures focusing on particular types of deviations can help to unravel post-transcriptional modification, maturation and degradation processes. Here, we focus on small RNA sequencing data that is produced in large quantities aimed at the analysis of microRNA expression. Starting from the recovery of many well known modified sites in tRNAs, we provide evidence that modified nucleotides are a pervasive phenomenon in these data sets. Regarding non-encoded nucleotides we concentrate on CCA tails, which surprisingly can be found in a diverse collection of transcripts including sub-populations of mature microRNAs. Although small RNA sequencing libraries alone are insufficient to obtain a complete picture, they can inform on many aspects of the complex processes of RNA maturation.

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