| Stripe rust, caused by Puccinia striiformis f. sp. tritici (Pst) is one of the most damaging diseases of wheat worldwide. Growing resistant cultivars is the most effective, economic and environment friendly way to control wheat pathogen and reduce the use of fungicide. Therefore, it is very important to investigate the resistant mechanism of the host plant and the pathogenic mechanism of the rust fungi the interaction between wheat and rust fungi in order to provide further information for selection and reasonable use of resistant wheat cultivars. In the previous study, the suppression subtractive hybridization (SSH) cDNA library of shuiyuanl 1 wheat leaves induced by Puccinia striiformis Chinese race CY23 was constructed, and more than 1000 ESTs was obtained. In present study, we generated the full length cDN A sequences of three genes from the SSH library. We also determined their expression in response to stripe rust pathogen infection and abiotic stresses. BSMV-VIGS (Barley Stripe Mosaic Virus-Virus Induced Gene Silencing) method was used for functional assessment of one gene TaLRPK. In addition, the resistant cultivars will become susceptible within a few years after release because the rapidly evolve of the rust pathogen. Therefore, it is essential to explore new genes from alien resources and related species of wheat in order to compete with the evolution of pathogen. We also searched for the new resistance from durum wheat and mapped it and transferred it into common wheat. The main results are as follows.1. A novel wheat gene, TaLRPK (Triticum aestivum leucine-rich-repeat receptor-like protein kinases), encoding a protein belonging to leucine-rich repeat receptor-like kinases (LRR-RLKs) was identified from wheat leaves infected by the stripe rust pathogen. It encodes 1045 amino acids with 42% of identity to Xa21 of Oryza longistaminata and contains a signal peptide, of 19 LRR motifs, a transmembrane domain, and a serine/threonine protein kinase domain. In transient assays with onion epidermal cells the TaLRPK-GFP fusion protein localized to the cytoplasm membrane. Transcripts were present at high levels in leaves, whereas low in other tested organs. The mRNA of TaLRPK was inducible in the compatible reaction between wheat leaves and stripe rust, and showed different profiles after treatment with salicylic acid, methyl jasmonate, ethylene, abscissic acid, wounding, low temperature and a salinity stress. Silencing TaLRPK using Barley Stripe Mosaic Virus-Induced Gene Silencing (BSMV-VIGS) reduced the transcript level of the gene, sporulation of the pathogen and development of hypersensitive response (HR) of the plants. These results suggest that TaLRPK may participate in the responses to environmental stresses and disease development in wheat.2. The methionine salvage pathway is conserved from prokaryotes and high eukaryotes. Aci-reductone dioxygenase (ARD) represent a branch point in the methionine salvage pathway. A novel aci-reductone-dioxygenase cDNA, designed as TaARD, was isolated from wheat leaves infected by the stripe rust pathogen. TaARD was predicted to encode a 197 amino-acid protein that belongs to the cupin superfamily. In transient assays with onion epidermal cells, the TaARD-GFP fusion protein localized to the nucleus and cytoplasm. Southern blot analysis showed that the wheat genome had multiple copies of TaARD. Quantity real time RT-PCR (qRT-PCR) analyses revealed that the TaARD transcript was induced in compatible interactions between wheat and the stripe rust pathogen. However, its expression was reduced or suppressed in incompatible interaction and by ABA, ethephon (ET), or salicylic acid (SA) treatments. Unlike the others, methyl jasmonate (MeJA) suppressed the TaARD expression in the first 6 hours but stimulated TaARD expression afterwards. The expression of TaARD also was inhibited by wounding and environmental stimuli, including high salinity and low temperature. These results suggest that TaARD may play a positive role in the metabolism of methionine and ethylene in response to biotic and abiotic stresses.3. A novel wheat gene, TaWRP (Triticum aestivum wound response protein), was identified in a subtraction library constructed with RNA isolated from wheat leaves infected with the stripe rust pathogen. It encodes 325 amino acids with 85% identity to wound response protein of Oryza sativa and contains unknown function DUF151 domain. The transcripts were present at high levels in leaves, whereas low in other tested organs. The mRNA of TaWRP was inducible in the incompatible interaction or by SA or by wounding, while suppressed in the compatible interaction or by ET, ABA, JA or by high salinity stress. The results suggest that TaWRP is a wound response protein, which may be regulated by the SA pathway, and may participate in the responses to environmental stresses and disease resistance in wheat.4. Durum wheat germplasm has excellent resistance to stripe rust, but not many genes for stripe rust resistance had been identified from durum wheat genotypes. Durum wheat PI 480148, originally from Ethiopia, was resistant in all seedling tests with several US races under controlled greenhouse conditions and at multiple locations under natural infection of the pathogen for four years. To map the resistance gene(s) in the genotype and transfer it into common wheat, a cross was made between PI 480148 and susceptible common wheat genotype Avocet Susceptible (AVS). Resistant F3 plants with 42 chromosomes were selected through Feulgen-staining of root tip cells and testing with Pst races. When tested with PST-100, the most predominant race in the US for the last six years,157 F4 plants from a single F3 hexaploid plant segregated into a 3:1 ratio for resistant to susceptible plants, which identified a single dominant gene from PI 480148. Using the F3:4 population and the resistance gene-analog polymorphism (RGAP) and simple sequence repeat (SSR) techniques, the gene was mapped to the long arm of chromosome 2B. An RGAP marker and a SSR marker (Xwmc441) were closely linked to the resistance gene with a genetic distance of 2.7 and 5.6 cM, respectively. The effective resistance of the gene to an Australian isolate virulent to Yr5, which is located on 2BL and resistant to all Pst races in the US so far, indicated that the gene is different from Yr5 and should be a new and useful gene for resistance to stripe rust. Resistant common wheat lines with plant types similar to AVS were selected for use in breeding programs to develop common wheat cultivars with the resistance gene. This approach is currently used to identify and transfer different genes for effective resistance from a number of durum wheat genotypes to a common wheat background.
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