Resistance to powdery mildew in wheat germplasm with different resistance sources L. M. Miranda, J. P. Murphy, D. S. Marshall and S.Leath NC STATE UNIVERSITY Contact information: METHODOLOGY  Powdery mildew resistant lines (Table 1) NC96BGTD3 (NCD3), NC96BGTA5 (NCA5), NC96BGTA6 (NCA6) and NC97BGTD7 (NCD7) were each crossed to the susceptible cultivar ‘Saluda’  F2 derived lines (90-120) from these four populations were evaluated for their disease reaction in greenhouse and field experiments following inoculation and rating procedures described by Srnic et al. (2005).  Germplasm x germplasm populations: NCD3xNCD7, NCA6xNCD7, NCA5xNCD7 and NCA5xNC99BGTAG11(NCAG11) were also rated in the field (Srnic et al identified SSR markers for NCAG11)  DNA extractions followed method by Stein et al. (2001) and PCR protocols for the linkage analysis with SSR markers were performed as described by Srnic et al.(2005).  Virulence test followed procedure by Leath and Heun (1990) with powdery mildew isolates provided by R. Parks and C. Cowger (USDA-ARS Raleigh, NC) INTRODUCTION Powdery mildew of wheat, caused by Blumeria graminis f.sp. tritici, is a prevalent fungal disease in the southeastern United States. Host resistance as a method to control the disease requires plant breeders to constantly search for new resistance sources. The North Carolina State University (NCSU) Small Grains Programs has released 11 germplasm lines with powdery mildew resistance introgressed from diploid and tetraploid wheat relatives into the soft red winter wheat cultivar ‘Saluda’. Molecular markers  SSR markers linked to the four powdery mildew resistance genes were found.  Linkage maps were constructed (Figures 2 and 3) Table 2. Differential response of NC D3, NC D7, Ulka (Pm2) and Chancellor (no Pm genes) after inoculation with five isolates of B. graminis f.sp. Tritici based on rating scale by Leath and Heun (1990) Isolate Line/cultivar NCD3NCD7Ulka(Pm2)Chancellor H C E J J Candidate new Pm genes  Pm genes in NC lines were putatively assigned to specific chromosome arms based on the chromosomal location of the SSR markers linked to them but not all SSR markers found are reported to be chromosome specific in the literature.  Only NCD7 and NCD3 genes are in a chromosome arm were no order Pm gene has been reported (5DL).  Pm 2 is also located in chromosome 5D but in the short arm.  Virulence test differentiated among disease reactions of NCD7, NCD3 and Ulka (Pm2) (Table 2, Figure 4)  Chromosomal location of all SSR markers linked to the NCD7 Pm resistance was confirmed using Nullitetrasomic (Nulli5D-tetra5A) and ditelosomic 5DL (DT5DL) ‘Chinese spring’ lines:  SSR marker bands for Xbarc144, Xbarc177 and Xgwm 272 were absent in Nullisomic 5D but present in DT5DL and in Chinese spring (Figure 5)  Additional markers are needed to confirm chromosomal location of NCD3 gene. Germplasm x germplasm populations  Codominant SSR markers were used to selected for homozygous resistant lines among those phenotypically scored as resistant. NCA5 Pm resistance Xgdm33 Cfa2153 NCA6 Xbarc121 Cfa2019 Cfa2123 Pm resistance cM Chromosome1AS Chromosome 7AL NCD7 Xbarc144 Xgwm272 Xbarc177 Pm resistance NCD3 Pm resistance Cfd cM Chromosome 5DL OBJECTIVES  To determine the mode of inheritance of powdery mildew resistance in four wheat germplasm lines developed by NCSU Small Grains Program (Table 1)  To identify microsatellite (SSR) markers linked to these powdery mildew resistance genes that can be used for Marker Assisted Selection (MAS). Germplasm LineSource of resistance NC96BGTA5 (NCA5)T. monococcum subsp. aegilopoides NC96BGTA6 (NCA6)T. monococcum subsp. aegilopoides NC96BGTD3 (NCD3)Ae. tauschii subsp. strangulata NC97BGTD7 (NCD7)Ae. tauschii Table 1. Germplasm lines and their source of resistance to powdery mildew Figure 1. Comparison of field reaction to powdery mildew between North Carolina germplasm line NCA6 and susceptible cultivar ‘Saluda’ NCA6Saluda Figure 2. Linkage maps for powdery mildew (Pm) resistance in NCA5 and NCA6 constructed with SSR markers using MAPMAKER Exp Version 3.0b (Kosambi mapping function, LOD 3.0) Figure 3. Linkage maps for powdery mildew (Pm) resistance in NCD7 and NCD3 constructed with SSR markers using MAPMAKER Exp Version 3.0b (Kosambi mapping function, LOD 3.0) Figure 5. Chromosomal localization of SSR marker Xbarc177 NC D7 Saluda Ae.Tauschii C.S. N5D DT5DL 137.8bp 128.8bp 148bp PCR products observed in NCD7, Saluda, Ae. Tauschii, Chinese Spring (C.S.) and Ditelosomic 5DL (DT5DL) but no PCR product observed in Nullisomic 5D (N5D). Similar results observed with Xbarc144 and Xgwm 272 (images not shown) Results Phenotypic analysis  Greenhouse and field disease ratings (Figure 1) showed segregation for powdery mildew resistance as a monogenic dominant trait for all germplasm lines (X 2 tests for 1 resistant:2 segregating:1 susceptible ratio were non significant, P>0.46).  Germplasm x germplasm populations fitted a digenic model except for NCD3xNCD7. Isolate H2-3 NC D3 NCD7 Ulka Chancellor NC D3 NCD7 Ulka Chancellor Isolate E2-5 Figure 4. Virulence test for NCD3, NCD7, Ulka (Pm2) and Chancellor using B. graminis f.sp. tritici isolates E2-5 and H2-3 Conclusions  Powdery mildew resistance is inherited as a monogenic dominant trait in the four NC germplasm lines included in the study.  SSR markers found can be used to be develop lines with pyramids of Pm genes or to facilitate the incorporation of these new resistance sources into cultivar development programs.  The powdery mildew resistance gene in NCD7 is postulated as a new Pm gene. Literature cited Leath, S and Heun, M Identification of powdery mildew resistance genes in cultivars of soft red winter wheat. Plant disease 74: Srnic, G., Murphy, J.P., Lyerly, J.H., Leath, S. and Marshall, D.S Inheritance and chromosomal assignment of powdery mildew resistance genes in two winter wheat germplasm lines. Crop Science 45: Stein, N., Herren, G. and Keller, B A new DNA extraction method for high-throughput marker analysis in a large genome species such as Triticum aestivum. 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