2012 Annual Report 1a.Objectives (from AD-416): Conduct research to identify, characterize, and determine the function of genes involved in nitrogen fixation, nitrogen assimilation, phosphorus acquisition, and root development in Lupinus, Medicago, and Phaseolus. 1b.Approach (from AD-416): 1. Prepare cDNA libraries from various plant organs of the three legume species. Develop genomic libraries of the three species. 2. Use high throughput sequencing to evaluate transcriptome. 3. Sequence genes and prepare antibodies to the gene products. 4. Evaluate gene expression via RNA blots, immunoblots, enzyme assays, and in situ hybridization. 5. Characterize potential promoter elements for important genes related to the processes cited above. 6. Modify expression of relevant genes via overexpression, antisense, and/or RNAi approaches. BSL-1; Re-Certified June 12, 2007. 3.Progress Report: The research reported in this project addresses Objective 2 of the parent project 3640-21000-028-00D: Develop and employ RNA interference-mediated gene silencing to identify the functional role of genes involved in phosphorus and nitrogen acquisition and metabolism in root tissues of legumes such as common bean, pea, and lentil. Phosphorus (P) is a non-renewable fertilizer resource required for plant growth and development. Easily mined P is expected to peak between 2040 and 2060. Developing crop plants that are more efficient at acquiring and using P is critical for world agriculture. To address plant genes responsive to and regulating P acquisition and use, ARS researchers in St. Paul, Minnesota, in cooperation with collaborators at the University of Minnesota, performed high-throughput transcript sequencing (RNA-seq) on white lupin plants that were grown under either phosphorus-deficient or phosphorus-sufficient conditions. We discovered more than 2,000 genes that were regulated by phosphorus. This experiment showed that phosphorus-deficient plants had increased abundance of RNA for plant growth hormones, polyamine production, and one carbon metabolism. We identified 12 genes that could be useful in marker-assisted plant breeding to improve phosphorus acquisition in plants.