Application of Parallel Reaction Monitoring in 15N Labeled Samples for Quantification

Accurate relative quantification is critical in proteomic studies. The incorporation of stable isotope 15 N to plant-expressed proteins in vivo is a powerful tool for accurate quantification with a major advantage of reducing preparative and analytical variabilities. However, 15 N labeling quantific...

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Bibliographic Details
Published in:Frontiers in plant science 2022-05, Vol.13, p.832585-832585
Main Authors: Reyes, Andres V., Shrestha, Ruben, Baker, Peter R., Chalkley, Robert J., Xu, Shou-Ling
Format: Article
Language:English
Subjects:
15N metabolic labeling
parallel reaction monitoring
Plant Science
proteomics
skyline
targeted quantification
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Summary:Accurate relative quantification is critical in proteomic studies. The incorporation of stable isotope 15 N to plant-expressed proteins in vivo is a powerful tool for accurate quantification with a major advantage of reducing preparative and analytical variabilities. However, 15 N labeling quantification has several challenges. Less identifications are often observed in the heavy-labeled samples because of incomplete labeling, resulting in missing values in reciprocal labeling experiments. Inaccurate quantification can happen when there is contamination from co-eluting peptides or chemical noise in the MS 1 survey scan. These drawbacks in quantification can be more pronounced in less abundant but biologically interesting proteins, which often have very few identified peptides. Here, we demonstrate the application of parallel reaction monitoring (PRM) to 15 N labeled samples on a high resolution, high mass accuracy Orbitrap mass spectrometer to achieve reliable quantification even of low abundance proteins in samples.
ISSN:1664-462X
1664-462X
DOI:10.3389/fpls.2022.832585