LC-MS/MS analysis of an intact synthetic microRNA

The field of epitranscriptomics is growing in importance, with chemical modification of RNA being associated with a wide variety of biological phenomena. Mass spectrometry (MS) enables the identification of modified RNA residues within their sequence contexts, by using analogous approaches to shotgun proteomics. We have developed a free and open-source database search engine for RNA MS data, called NucleicAcidSearchEngine (NASE), as part of the OpenMS software framework. NASE allows the reliable identification of (modified) RNA sequences from LC-MS/MS data in a high-throughput fashion. For this validation dataset, oligonucleotides with the sequence of mature Drosophila let-7 microRNA, 21 nt in length, were produced synthetically in unmodified and modified (2’-O-methylated at the 3’ uridine) forms. We characterised a 1:1 mixture of both forms of this RNA.