Metabolic Profiling Allows Comprehensive Phenotyping of Genetically or Environmentally Modified Plant Systems

Abstract

Metabolic profiling using gas chromatography-mass spectrometry technologies is a technique whose potential in the field of functional genomics is largely untapped. To demonstrate the general usefulness of this technique, we applied to diverse plant genotypes a recently developed profiling protocol that allows detection of a wide range of hydrophilic metabolites within a single chromatographic run. For this purpose, we chose four independent potato genotypes characterized by modifications in sucrose metabolism. Using data-mining tools, including hierarchical cluster analysis and principle component analysis, we were able to assign clusters to the individual plant systems and to determine relative distances between these clusters. Extraction analysis allowed identification of the most important components of these clusters. Furthermore, correlation analysis revealed close linkages between a broad spectrum of metabolites. In a second, complementary approach, we subjected wild-type potato tissue to environmental manipulations. The metabolic profiles from these experiments were compared with the data sets obtained for the transgenic systems, thus illustrating the potential of metabolic profiling in assessing how a genetic modification can be phenocopied by environmental conditions. In summary, these data demonstrate the use of metabolic profiling in conjunction with data-mining tools as a technique for the comprehensive characterization of a plant genotype.

Varietal characterization of hop (Humulus lupulus L.) by GC–MS analysis of hop cone extracts

“…Derivatization was performed according to recognized protocols [8]. Dried extracts were reconstituted in 10 mL of a 40 mg/mL solution of methoxyamine hydrochloride in pyridine and heated for 90 min at 301C.…”

“…Other investigators have differentiated hop varieties by using analysis of hop acids or flavonoids using reversed phase HPLC [2]. In 2001 Roessner et al showed that profiling metabolites in plant extracts allows comprehensive phenotyping of genetically or environmentally modified plant systems [8]. Such studies draw on simple extraction procedures that have been shown to be very robust [9] and have permitted wide-ranging high-throughput applications, such as phenotyping to diagnostic analyses in plants.…”

Metabolomics — the link between genotypes and phenotypes

“…Instead of losing information in the process of averaging metabolite levels, each metabolite profile represents a true and valid response of metabolism upon subtle (but unknown) changes in parameters of the system. By computing correlation coefficients of metabolite-metabolite plots, the detection of homeostatic regulation of metabolite ratios could be set onto a more rigid statistical basis (Arkin et al, 1997;Roessner et al, 2001;Kose et al, 2001). A computation of metabolic models based on a combination of co-response MCA theory and experimentally detected metabolomic correlations, however, remains to be shown.…”

“…If this is not the case, the data acquisition might have been obscured by either sample preparation faults or low analytical precision during data acquisition. Eventually, subtle mutations that result in silent phenotypes might not render a clear clustering result by any common means such as principal component analysis (Fiehn et al, 2000b;Roessner et al, 2001) or discriminatory analysis (Raamsdonk et al, 2001). For such cases, further methods need to be developed that unravel potential differences between the populations being studied.…”

The cell death factor, cell wall elicitor of rice blast fungus (Magnaporthe grisea) causes metabolic alterations including GABA shunt in rice cultured cells

“…Recently, analytical methods such as liquid chromatography-mass spectrometry, gas chromatography-mass spectrometry, nuclear magnetic resonance and capillary electrophoresis-mass spectrometry (CE/MS) provided new insights into the biological function of plant metabolism. [19][20][21][22] Here, using capillary electrophoresis-mass spectrometry (CE/MS), we determined the concentrations of target metabolites in rice suspension cells treated with M. grisea elicitor to facilitate understanding in the early steps of elicitor-induced processes. Overall changes in the metabolite levels in the central pathways were observed.…”