2010 Annual Report 1a.Objectives (from AD-416) 1) To develop soybeans with altered seed coat color to facilitate the segregation and identity preservation of seeds with genetically enhanced compositional traits.. 2)To produce soybeans with enhanced food, feed, and industrial properties by modification of the oil and lipid-soluble antioxidant composition of seeds.. 3)To introduce genes into soybeans that result in high-level accumulation of foreign and engineered proteins valuable for food, feed, and industrial uses.. 4)To develop procedures and methods for analyzing and predicting seed protein allergenicity in food and feed. To develop non-allergenic seeds by suppressing intrinsic allergens and to modify proteins that are potential transgenes for improving biosafety. 1b.Approach (from AD-416) 1) Co-transformation of soybean with a seed coat color-conferring transgene and a trait transgene will yield visually distinct seeds with enhanced composition.. 2)Identify enzymes that are specialized for the metabolism of unusual fatty acids to produce agronomically-viable soybean seeds.. 3)Manipulate protein content of soybean seeds by use of plant promoters and compartmentalization of introduced proteins.. 4)Identify and characterize soybean food and feed allergens. 3.Progress Report This project is focused on translating genomics and associated processes for the improvement of a major field crop, soybean. The parent project plan for this research focus was discarded upon the retirement of the Lead Scientist and has since been redirected to encompass the NPS approved research focus of two new scientists, one appointed in late FY09 and the other in early FY10. Preliminary research objectives have been established and approved for one of the scientists and objectives for the other are under development. A combined project plan for Agency evaluation is currently in preparation. The research objectives required the establishment of a high throughput element analysis facility to assay the elemental composition (ionome) of plant tissues, in particular seeds. This facility is now on line and operational, a process that took several months. Several collaborations with ARS scientists interested in soybean seed quality traits and their relationship to elemental profiles have been initiated and the research is underway. The elemental composition of second generation seeds from crosses of low and high phytic acid (an anti nutrient that limits available P and other elements) has been completed. A second study into the effect of soil environments on element accumulation in the seed using publically available data has been completed and submitted for publication. The results of this study will form the basis of a system approach to understanding the gene X environment interactions governing seed composition. An in vitro soybean embryo culture system has been developed and experiments to evaluate its ability to mimic natural seed conditions are in progress. Metabolite labeling studies aimed at understanding sub cellular compartmentalization of metabolism have been initiated and data analysis is in progress. 4.Accomplishments 1. Environmental control of seed elemental composition. Understanding how plants regulate element composition of tissues is critical for agriculture, the environment, and human health. Sustainably meeting the increasing food and biofuel demands of the planet will require growing crops with fewer inputs such as the primary macronutrients phosphorus (P) and potassium (K). Ionomics is the study of elemental accumulation in living systems using high-throughput elemental profiling. With this technique, large quantities of data on thousands of samples can be generated. Scientists at the Plant Genetics Research Unit in St. Louis, MO used this approach to sample the natural diversity present in collections of a model plant, the wild mustard Arabidopsis, and mapping populations derived from those collections. The data that was generated was used in a genetic mapping protocol to uncover hundreds of genetic loci important for elemental accumulation in the seeds of this model plant. It was discovered that when the soil environment changes, the pattern of genes throughout the genome that are important for elemental accumulation change in a corresponding fashion. In addition, the elements themselves have different inter-relationships depending on the environment and the lines under study. For example, in one soil environment two elements may accumulate independently from one another where in a different environment they co-accumulate. The findings suggest that crop varieties that are developed for improved elemental uptake and accumulation will be highly environment specific. This finding has important ramifications for breeders concerned with plant nutrition (elemental uptake from the soil) and yield as well as the quality and composition of the seeds. Improving the quality of the elemental profiles of seeds is important for human nutrition and animal feed. 2. Sub-cellular compartmentalization of metabolism. A current limitation to improving seed quality and composition using biotechnology is a lack of understanding of how compounds are partitioned within the cells of seed tissues and how carbon and nitrogen from the parent plant flow into the various compartments. One effective means of addressing this problem is to take a Systems Biology approach where an overall picture of metabolism within the seed can be addressed through time and space (cellular and tissue). This approach is limited in plants by a lack of experimental data that maintains spatial integrity and reports on organelle-level metabolism. Scientists of the Plant Genetics Research Unit in St. Louis, MO used cultured embryos of rapeseed, as a prelude to soybean, along with isotopic labels, including 13Carbon and 2Hydrogen to analyze metabolism at the subcellular level as a means of addressing this problem. Gas chromatography- Mass Spectrometry (GCMS) and Nuclear Magnetic Resonance (NMR) were employed to assess compartmentalization in two ways, amino acid labeling in protein subunits that are synthesized in different organelles, and [U-13C]-compounds for organelle partitioning of basic metabolism. Preliminary data suggests that this approach will generate the necessary data to measure the flux of metabolites into subcellular compartments. The data that is generated will allow for the development of strategies, either biotechnological or conventional, to engineer seeds with improved characteristics for quality and compositional traits, an important goal for soybean producers in the U.S. Review Publications Morrissey, J., Baxter, I.R., Lee, J., Li, L., Lahner, B., Grotz, N., Kaplan, J., Salt, D.E., Guerinot, M. 2009. The Ferroportin Metal Efflux Proteins Function in Iron and Cobalt Homeostasis in Arabidopsis. The Plant Cell. 21:3326-3338. Baxter, I.R. 2010. Ionomics. Briefings in Functional Genomics and Proteomics. 9:149-156. Vogel, J.P., Garvin, D.F., Gu, Y.Q., Lazo, G.R., Anderson, O.D., Bragg, J.N., Chingcuanco, D.L., Weng, Y., Belknap, W.R., Thomson, J.G., Dardick, C.D., Baxter, I.R. 2010. Genome sequencing and analysis of the model grass Brachypodium distachyon. Nature. 463:763-768.