2006 Annual Report 1.What major problem or issue is being resolved and how are you resolving it (summarize project aims and objectives)? How serious is the problem? Why does it matter? Asian soybean rust (ASR), caused by Phakopsora pachyrhizi, is the most threatening emerging disease for the U.S. soybean crop. None of the 18,000 soybean accessions from the USDA germplasm collection evaluated in seedling assays has shown genetic resistance to all isolates. A few accessions have genetic resistance to one isolate and one accession has some resistance or tolerance to four of twelve isolates examined. In light of the obvious vulnerability, soybean rust could cause 50% crop losses and have an economic impact of $7B/yr on U.S. soybean production. Little is known about the infection process. Molecular aspects of resistance and susceptibility of soybean to rust have not been studied. This knowledge is needed to develop new modes of resistance using biotechnology, because present germplasm resources likely will not provide resistance to most rust isolates. The soybean cyst nematode (SCN), Heterodera glycines, is currently the most economically destructive pathogen of soybean and causes an estimated loss of approximately one billion dollars per year in the U.S. Currently, there are very few genetic loci for resistance to SCN. These resistance loci do not provide protection against all SCN isolates; therefore breeders cannot protect our soybean crop by pyramiding only naturally occurring resistance known to exist in soybean. A foundation of knowledge is lacking at the molecular and cellular levels regarding the infection process and the response of susceptible and resistant soybean plants to invasion by SCN. This knowledge will be useful to design new modes of resistance to SCN using biotechnology to insure that our soybean crop is protected as new strains of SCN evolve. Furthermore, there are other important threats to soybean that warrant the development of broader resistance of soybean to pests and pathogens and which require the development of a foundation of knowledge at the molecular and cellular levels. The project has three specific objectives:. 1)Discover and characterize plant and pathogen genes important for resistance or pathogenicity at the molecular level with special emphasis on, but not limited to soybean interactions with soybean rust and soybean cyst nematode; . 2)Determine modes of action for plant disease resistance genes, pathogen virulence factors and molecular signals responsible for host-parasite interactions through analysis and characterization of genetic, molecular, protein and metabolite networks;. 3) Engineer and evaluate new methods for obtaining resistance, such as gene silencing, over-expression, protein antagonism, and chemical or biological inhibition of host and pathogen processes, with special emphasis on soybean rust and the soybean cyst nematode. The research to be undertaken falls under National Program 302 - Plant Biological and Molecular Processes and addresses component 1 and 2. Specifically these are: 2.2.1.2 Pathogen Interactions with Host Plants and 2.1.2.3 Plant Genome Mapping. The research will result in the discovery of fundamental information leading a better understanding of plant-microbe interactions and to improved resistance of soybean and other crop plants to pests and pathogens and will result in technology and molecular tools leading to broader disease resistance of soybean and other crop plants. The molecular tools developed as a result of this project will be used by scientists studying plant/disease interactions and by soybean breeders to develop SCN resistant cultivars. Broadening resistance of soybean-to-soybean rust, soybean cyst nematode, and other pests and pathogens will reduce levels of pesticide applied to crops that enter into the environment. Furthermore, increased yield will translate into reduced costs to producers, users and consumers. 2.List by year the currently approved milestones (indicators of research progress) This is a new project and was only recently approved to replace project 1275-21220-214-00D. The new milestones are as follows: Year 1 (FY 2007) a) Obtain tissue, sections, and isolate cells for cDNA libraries and protein analysis. b) Collect susceptible and resistant tissues. c) Identify up-regulated cell wall hydrolases. Year 2 (FY 2008) a) Extract RNA and proteins, construct libraries and analyze protein. b) Prepare and hybridize probe to microarrays. c) Isolate gene promoters for selected up-regulated hydrolase genes. d) Prepare yeast 2-hybrid libraries from pathogen infected tissues. e) Isolate full-length cDNA of candidate genes. Year 3 (FY 2009) a) One-pass sequence 1,000 to 3,000 clones. b) Annotate proteins. c) Put data on-line. d) GUS/GFP analysis of cell wall hydrolase genes. e) RT-PCR analysis of microarray results. f) Compare microarray and proteomics results in soybean to defense response networks in Arabidopsis. g) Construct over-expression vector with candidate defense genes. h) Prepare RNAi constructs with infection-targeted promoters. Year 4 (FY 2010) a) Submit to GenBank one-pass sequence from both ends of 5,000 clones from each library. b) GUS/GFP expression analysis of up-regulated genes. c) Identify common components in signaling pathways in Arabidopsis. d) Transform soybean roots and infect with SCN. Year 5 (2011) a) Annotate ESTs. b) GUS/GFP analysis of up-regulated genes. c) Assemble defense and signaling pathways. d) Analyze transformed roots. e) Transform and regenerate soybean plants for further analysis. 4a.List the single most significant research accomplishment during FY 2006. This project is a new and was only recently approved to replace project 1275-21220-214-00D. Details of accomplishments are in annual report for project 1275-21220-214-00D. 4c.List significant activities that support special target populations. NONE 5.Describe the major accomplishments to date and their predicted or actual impact. This project is new and was only recently approved to replace project 1275-21220-214-00D. Details of accomplishments are in annual report for project 1275-21220-214-00D. 6.What science and/or technologies have been transferred and to whom? When is the science and/or technology likely to become available to the end-user (industry, farmer, other scientists)? What are the constraints, if known, to the adoption and durability of the technology products? This project is new and was only recently approved to replace project 1275-21220-214-00D. Details of science and/or technologies transferred are in the annual report for project 1275-21220-214-00D. 7.List your most important publications in the popular press and presentations to organizations and articles written about your work. (NOTE: List your peer reviewed publications below). This project is new and was only recently approved to replace project 1275-21220-214-00D. See project 1275-21220-214-00D for FY 2006 information.