Currently, interest is surging over the discovery of techniques that can identify and manipulate the molecular basis of macroscopic plant phenotypes. Metabolic flux analysis, measuring the flow of matter through an organism’s metabolic pathways, is one of the best approaches to resolve the intricate relationship between molecular composition and expressed traits. The research displayed here seeks to develop robust in vivo isotopic labeling techniques for improving metabolic flux analysis.
In vivo isotopic labeling of plant systems is accomplished by the stable (nonradioactive) 12C and 13C isotopologues. Since carbon atoms are present in virtually all metabolites and cellular macromolecules, 13C introduced via 13CO2 gas is readily assimilated into the plant metabolic systems and high enrichment is achieved. Metabolic flux is measured by varying the isotopologue composition in the environment during growth and analyzing the turnover of metabolites in tissues by mass spectrometry.
Labeling of live plants is performed in an isotopic growth chamber that provides a well regulated 13CO2 environment. Due to the high cost of the isotope, all 13CO2 labeling techniques utilize an enclosed environment which minimizes 13CO2 gas loss and 12CO2 gas contamination. However, growing organisms in an isolated environment raises many additional challenges concerning the availability of nutrients and the buildup of metabolic products. The nature of 13CO2 labeling requires that viable growth conditions be maintained in a way that does not compromise the integrity of the 13C labeling composition. Therefore, all growth chambers implement automation to reliably monitor and regulate stresses, ensuring optimal conditions for high enrichment 13C labeling.