1 Metabolic phenotyping of genetically diverged species in

Systematic evaluation of phenotypic variations among cereal breeds based on metabolic profiling is an effective approach to holistic discovery of metabolic markers as well as nutrition targeted breeding. Furthermore, integrated analysis of metabolo-phenotype and genotype of natural variations or cultivars should be useful to find out metabolic markers in association with genotypes. We have applied NMR methods to perform metabolic profiling in Gramineae plants (wheat, barley and rice), in order to develop a novel metabolic-phenotyping procedure by global measurement of metabolite in various samples. Solution samples extracted from seed grains of 21 wheat, 21 barley and 18 rice strains were conducted for the metabolic profiling by 1D- 1H-NMR and 2D-1H-1H NMR spectra. The metabolic fingerprint of each of 60 strains by the 1D-1H-NMR were analysed using principal component analysis (PCA) and hierarchical clustering analysis (HCA) to compare metabolic profiling among species and to find out metabolic marker by chemical shift values corresponding to metabolite specifically abundant in each strain. Furthermore, 2D-1H-1H NMR spectral profiling was also applied to quantify major metabolites. In total, 22 metabolites were compared among strains by the 2D-NMR spectra. These major metabolite profiles allow us to evaluate nutritious balance in cereal grains. Furthermore, in order to compare genetic divergence and metabolo-phenotypic variations, we performed genome wide genotyping using AFLP analysis for comparison between genotype and metbolo-phenotype to discover metabolites in association with genetic polymorphisms. NMR based metabolic-phenotyping and that integration with genome wide genotyping should be mostly applicable to systematic exploration of cereal genetic resources as well as to metabolite based breeding for enhancement of cereal productivity