Analytical Methods for Characterizing RNA by LC-MS/MS

The Central Dogma of Molecular Biology states the genetic information stored in our DNA is transcribed into an intermediate molecule, RNA, which is then translated into protein. There are three main types of RNA associated with transcription/translation: messenger RNA, ribosomal RNA, and transfer RNA. All RNAs in the biological system carry post transcriptional modifications: chemical functional groups added to the base, the ribose sugar, or both. The gold standard in analytical technology for identification and characterization of these modifications remains steadfast in the realm of liquid chromatography hyphenated to high resolution accurate mass spectrometry. This paper covers how the analysis of native RNA is technically demanding and how RP-based separation strategies that avoid ion-pairing reagents are in demand. It highlights the use of Orbitrap Tribid MS for system evaluation with and without sulfolane to assess retention, signal stability, and fragmentation efficiency.