Poster abstracts
Poster number 118 submitted by Asif Rayhan
Nucleoside modification mapping by genome-independent universal mass exclusion list of unmodified oligonucleotides during LC-MS/MS
Asif Rayhan (Chemistry, University of Cincinnati), Balasubrahmanyam Addepall (Chemistry, University of Cincinnati), Patrick A. Limbach (Chemistry, University of Cincinnati)
Abstract:
To evaluate the effectiveness of the UMEL for mapping RNA modifications in a single organism, RNase T1 digestion products from different organisms tRNA were analyzed separately and the number of modified digestion products detected using standard DDA and UMEL were compared. However, modifications containing only pseudouridine or one dihydrouridine were not considered in these experiments as they have the same mass as uridine.
A standard DDA based LC-MS/MS analysis of RNase T1 digestion products from E.coli total tRNAs resulted in detection of 62 post-transcriptionally modified digestion products. When the
UMEL approach was used, total 72 post-transcriptionally modified digestion products were detected. Thus, the use of an exclusion list increased the number of detected modified digestion products by 16%. To demonstrate the applicability of the universal exclusion list for RNA modification mapping, S.pombe was chosen which is previously uncharacterized. The modified nucleosides in S.pombe have not been reported previously. That’s why a census of modification was obtained from LC-MS/MS analysis of a nucleoside digest. Seventeen unique modified nucleosides were identified from S.pombe tRNAs. The RNase T1 digestion products from S.pombe total tRNA were analyzed both the standard DDA and UMEL approaches. The UMEL approach identified 12% more post-transcriptionally modified digestion products than the standard DDA analysis. Notably, we observed improved data acquisition for modified oligonucleotides, as the instrument prioritizes time for modified species by excluding unmodified ones. We will extend our analysis to include Thrmus Themophillus and Citrobacter koseri.
Keywords: tRNA , LCMS