Poster abstracts
Poster number 1 submitted by Lauren Barnes
A Comparison of RNA Sequencing by Conventional and Nano Liquid Chromatography Tandem Mass Spectrometry Methods
Lauren E. Barnes (Department of Chemistry, University of Michigan, Ann Arbor, Michigan 48109, USA), Robert T. Kennedy (Department of Chemistry, University of Michigan, Ann Arbor, Michigan 48109, USA), Kristin S. Koutmou (Department of Chemistry, University of Michigan, Ann Arbor, Michigan 48109, USA)
Abstract:
Posttranscriptional chemical modifications of RNAs are crucial regulators of a number of cellular processes including protein synthesis. Despite the importance of these modifications, few methods exist to accurately assign the chemical identity and locations of RNA modifications. The gold standard for sequencing modified RNAs is high pressure liquid chromatography coupled to tandem mass spectrometry (HPLC-MS/MS). One of the major limitations of LC-MS/MS methods is the ionization of longer, and thus, sequentially unique, oligonucleotides. The most common setup is conventional liquid chromatography coupled to an MS/MS system with conventional electrospray ionization (ESI) as the source. Different ion sources have different ionization mechanisms and thus may provide different sequencing information. However, these differences have not been previously quantified for oligonucleotides. Here we compare HPLC-ESI-MS/MS to nanoLC-nanoESI-MS/MS to investigate their ionization efficiency and sequencing coverage of yeast tRNA prepared with two different enzyme digestions.
Keywords: RNA modifications, LC-MSMS, RNA sequencing