Barley yellow dwarf virus (BYDV), an aphid transmitted luteovirus, is the most widespread and economically damaging virus of cereal crops. The work in this thesis aims to characterise the basis of the naturally occurring resistance to BYDV in cereals in three ways: Firstly, by facilitating the isolation of the Yd2 gene for BYDV resistance from barley by a map-based approach. Secondly, by determining if a BYDV resistance gene in rice is orthologous to Yd2. Thirdly, by establishing if other BYDV resistance genes in non- Ethiopian barleys are allelic to Yd2. It is hoped that the information generated in this study will ultimately assist in the production of BYDV resistant cereal cultivars. A detailed genetic map of the Yd2 region of barley chromosome 3 was constructed, containing 19 RFLP loci, the centromere and the Yd2 gene. Yd2 mapped on the long arm, 0.5 cM from the centromere, and in the mapping population of 106 F2 individuals, perfectly cosegregated with the RFLP loci XYlp, and Xwg889. This map represents the first stage in a project to isolate the Yd2 gene by a map-based approach. The isolation of Yd2 could help to elucidate the molecular mechanism of the Yd2-mediated BYDV resistance, and may allow the production of BYDV resistant cereals by genetic transformation. The RFLP markers mapped closest to Yd2 could also be useful in barley breeding, by enabling selection for both the presence of Yd2 and the absence of agronomically undesirable traits known to be closely linked to Yd2. Genetically Directed Representational Difference Analysis (GDRDA) is a technique based on subtractive hybridisation, which can be used to identify RFLP markers closely linked to a gene of interest. Two GDRDA experiments were performed with the intention of generating additional RFLP markers close to Yd2. However, the first experiment yielded RFLP probes that were not derived from the barley genome, while the second experiment yielded probes that detected repetitive sequences. It was concluded that GDRDA is of limited use in generating further markers close to Yd2. To isolate the Yd2 gene by a map-based approach, a much larger mapping population will need to be analysed to genetically resolve markers tightly linked to Yd2. If the two morphological markers uzu dwarf and white stripe,,j flank Yd2, then they could assist in this task by enabling the visual identification of F2 seedlings resulting from recombination close to Yd2. However, in this study, both morphological markers were found to be located distal to Yd2. Therefore, these two morphological markers can not be used together to facilitate high resolution genetic mapping of the Yd2 locus. It may be possible to use large-insert genomic DNA clones from the relatively small genome of rice to generate further RFLP markers close to the Yd2 gene in barley, provided that the order of orthologous sequences in barley and rice is conserved close to the Yd2 locus. To assess the feasibility of this approach, RFLP probes used to identify loci close to Yd2 were mapped in rice using a segregating rice F2 population. Five of the RFLP loci mapped together and in the same order as RFLP loci mapped close to Yd2 in barley using the same probes. By comparing the location of RFLPs mapped by other researchers in rice using probes mapped close to Yd2, the region of conserved linkage between rice and the Yd2 region was tentatively identified as the central portion of rice chromosome 1. The collinearity shown by orthologous sequences in barley and rice indicated that it may indeed be possible to use rice to assist in generating RFLP markers close to Yd2. Of all the cereals, rice is the most amenable to map-based gene isolation, due to its small genome, well developed physical and genetic maps, and its ability to be genetically transformed with high efficiency. If a BYDV resistance gene that is orthologous to Yd2 could be identified in rice, this gene could be isolated with relative ease, and then used to identify barley cDNA clones corresponding to Yd2 gene by virtue of the sequence homology expected between these genes. To test if a BYDV resistance gene from an Italian rice line is orthologous to Yd2, recombinant-inbred rice lines previously characterised for this gene were analysed using probes mapped close to Yd2 in barley. No genetic linkage was detected between the RFLP loci and the BYDV resistance gene, indicating that the gene is unlikely to be orthologous to Yd2. BYDV resistance alleles at the Yd2 locus which are of a non-Ethiopian origin may show interesting differences to Ethiopian Yd2 resistance alleles. To identify barleys which may contain resistance alleles of Yd2, ten BYDV resistant barleys not known to contain Yd2 were assessed for their resistance to the PAVadel isolate of BYDV in the glasshouse. CI 1179, Rojo, Perry, Hannchen, Post and CI 4228 were found to be the most resistant under these conditions, and were analysed further. If the resistance from these barleys is controlled by alleles of Yd2, RFLP markers close to Yd2 will be expected to cosegregate with the resistance in F2 families derived from crosses between these resistant barleys and the BYDV susceptible barleys Atlas and Proctor. RFLPs suitable for use in these allelism tests were identified using probes mapped close to Yd2. However, time did not permit the analysis of these F2 populations. / Thesis (Ph.D.) -- University of Adelaide, Dept. of Plant Science, 1996
| Identifer | oai:union.ndltd.org:ADTP/264432 |
| Date | January 1996 |
| Creators | Collins, Nicholas C. |
| Source Sets | Australiasian Digital Theses Program |
| Detected Language | English |
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