title: LC-MS Analysis of Methylated RNA.
abstract: The detection and quantification of methylated RNA can be beneficial to understand certain cellular regulation processes such as transcriptional modulation of gene expression, immune response, or epigenetic alterations. Therefore, it is necessary to have methods available, which are extremely sensitive and accurate, for instance liquid chromatography-tandem mass spectrometry (LC-MS/MS). Here, we describe the preparation of RNA samples by enzymatic hydrolysis and the subsequent analysis of ribonucleosides by LC-MS/MS via NLS (Neutral loss scan) and DMRM (Dynamic multiple reaction monitoring). Also, we provide variations of these methods including chromatographic techniques and different kinds of quantification.
authors.affiliation: Institute of Pharmacy and Biochemistry, Johannes Gutenberg-University of Mainz, Staudingerweg 5, 55099, Mainz, Germany.
authors.forename: Kathrin
authors.initials: K
authors.lastname: Thüring
authors.affiliation: Institute of Pharmacy and Biochemistry, Johannes Gutenberg-University of Mainz, Staudingerweg 5, 55099, Mainz, Germany.
authors.forename: Katharina
authors.initials: K
authors.lastname: Schmid
authors.affiliation: Institute of Pharmacy and Biochemistry, Johannes Gutenberg-University of Mainz, Staudingerweg 5, 55099, Mainz, Germany.
authors.forename: Patrick
authors.initials: P
authors.lastname: Keller
authors.affiliation: Institute of Pharmacy and Biochemistry, Johannes Gutenberg-University of Mainz, Staudingerweg 5, 55099, Mainz, Germany. mhelm@uni-mainz.de.
authors.forename: Mark
authors.initials: M
authors.lastname: Helm
mesh_terms: D002138:Calibration; D002851:Chromatography, High Pressure Liquid; D002853:Chromatography, Liquid; D007553:Isotope Labeling; D008745:Methylation; D012313:RNA / Q000737:chemistry / Q000378:metabolism; D012260:Ribonucleases / Q000378:metabolism; D013223:Statistics as Topic / Q000379:methods; D053719:Tandem Mass Spectrometry; D057188:Workflow
chemical_list: D012313:RNA; D012260:Ribonucleases
keywords: DMRM; External calibration; LC-MS; NLS; RNA methylation; SIL-IS