2011 Annual Report 1a.Objectives (from AD-416) The primary objective of this cooperative research project is to rapidly deploy the rpg4/Rpg5/Rrf1 (rRR) resistance complex into agronomically advanced Midwestern malting barley cultivars. 1b.Approach (from AD-416) The Rpg5 locus and the region around ARD199 will be sequenced to quickly identify SNPs diagnostic for resistant and susceptible genotypes. A set of molecular markers flanking the rRR resistance complex will be developed so genotyping for the region can be performed by standard PCR or by using high-throughput technology. Marker-assisted selection (using these molecular markers) will then be used to rapidly deploy the rRR resistance complex in barley germplasm. Specifically, resistance will be introduced into the Ug99 susceptible two-rowed (Pinnacle and Conlon) and six-rowed (ND25160) elite malting barley lines. Preliminary disease screening will be conducted in Fargo, with followup screening each year at the BL3 facility in St. Paul and at the stem rust nursery in Kenya. 3.Progress Report The cloned rpg4/Rpg5 locus is the best source of resistance to the virulent stem rust race TTKSK (Ug99) in barley. The characterization of three different bi-parental populations determined that a third gene tightly linked to rpg4 and Rpg5, designated Rrf1, is also required for resistance to race Ug99. In the past year, three major discoveries have been made that clarify our understanding of the genetic interactions underlying the rpg4/Rpg5 locus:. 1)VIGS of each gene present at the Rpg5 locus determined that the rye stem rust resistance gene Rpg5 and a second unrelated NBS-LRR gene, HvRga1, are required together for resistance against race QCCJ and TTKSK.. 2)The two tightly linked NBS-LRR domain genes act in cooperation with a third genetically distinct yet tightly linked gene, Rrf1, to confer resistance. The Rrf1 gene has been delimited to a ~200 kbp region and three candidate Rrf1 genes have been identified.. 3)A protein phosphatase 2C (PP2C) gene at the locus may act as a dominant susceptibility factor determining the recessive nature of rpg4-mediated resistance. Molecular markers flanking the three genes required for resistance have been designed. The flanking markers are diagnostic of the entire rpg4/Rpg5/Rrf1 (rRR) locus and we are currently using the LRK1 marker designed from the Rpg5 sequence to identify homozygous rRR positive BC1 F2 backcross lines. Marker-assisted selection using the molecular marker LRK1 is being utilized to rapidly deploy the rRR resistance complex. The rRR complex was introduced into Ug99 susceptible two-rowed and six-rowed elite malting barley lines. The two-rowed and six-rowed near isogenic lines with genetically defined rRR regions were used as the donor parents and the initial crosses were made in 2010. Currently, BC1F1 plants are growing in the growth chamber for BC1F2 seed production to continue backcrossing to BC5. This research has had an immediate impact on the rapid deployment of Ug99 resistance into elite two and six-rowed malting barleys adapted to the Midwest. In the next year, pyramiding the rRR resistance locus with a new source of resistance identified in the barley landrace designated Sw645 will begin and the BC3 lines containing the rRR locus will be sent for Ug99 evaluation in the Kenya stem rust nursery. We expect to have BC5 breeding lines with the rRR locus available by the 2013 growing season and are working to use SNP marker technologies to map a new source of resistance to pyramid with this locus. The three way crosses have been made and we plan to develop the molecular markers to the new gene in the coming year. We will also use SNP technology to screen the background of our BC2F1s to select lines with low percent donor parent genome to rapidly recover the recipient genome. ADODR monitoring activities to evaluate research progress included face-to-face meetings and discussions between project participants during the fiscal year.