2009 Annual Report 1a.Objectives (from AD-416) 1) Correlate metabolites during seed development with final seed composition; and. 2)Explore the basis for variation in seed composition as a function of canopy position. 1b.Approach (from AD-416) The proposed studies will focus on three broad aims. First, we will build on our observation that the free pool of amino acids (in particular asparagine) during development is correlated positively with seed protein content at maturity. Free amino acids will be extracted from rapidly growing seeds and quantitated using GC-MS. We will attempt to identify quantitative trait loci associated with the free asparagine pool in developing seeds of a segregating inbred population that has previously been genotyped. Concurrent studies will examine the levels of enzymes involved in asparagine metabolism (asparaginase and asparagine synthetase) using custom anti-peptide antibodies for immunoblotting analysis. Second, we will initiate a new effort to identify soybean genotypes containing high levels of sulfur amino acids (methionine and cysteine) in mature seeds. If preliminary findings can be confirmed, appropriate crosses will be made to begin to move the high-sulfur trait into enhanced germplasm lines. Third, we will continue to examine the effect of canopy position on seed composition. In general, seeds that develop at the top of the canopy have more protein, less oil, and fatty acids contain more monounsaturated oleic (18:1), compared to seeds at the bottom of the canopy. Our current results suggest that quantitative differences exist among genotypes in these responses. In the proposed studies, we will determine whether these canopy position effects may simply reflect sensitivity to environmental parameters within the canopy and concurrently, appropriate crosses will be made to identify controlling genes. 3.Progress Report The overall goal of this research is to identify physiological determinants that control or impact seed composition and a major focus is on the variation in seed composition as a function of canopy position. Seeds produced at the top of the canopy tend to be higher in protein and lower in oil, and the oil has a higher percentage of the desirable fatty acid, oleic acid (18:1). These top-to-bottom gradients in seed composition within the canopy are consistently observed but differ quantitatively among genotypes. In order to determine the extent to which differences reflect inherent physiological features of the mother plant as opposed to micro-environment within the canopy, plants in the field were thinned at flowering from the normal dense stands to single plants to allow for more uniform environmental conditions along the main stem. The preliminary results indicate that the thinning treatment did not substantially affect the canopy variation in seed oil conent, suggesting that genetic factors outweigh environmental factors. Collectively, these studies may help to develop soybean cultivars with increased seed protein content and greater compositional stability.