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Towards map-based cloning of Fusarium head blight resistance QTL Fhb1 and non-additive expression of homoeologous genes in allohexaploid wheatPumphrey, Michael Odell January 1900 (has links)
Doctor of Philosophy / Department of Plant Pathology / Bikram S. Gill / Wheat is the most widely grown and consumed grain crop in the world. In order to meet future agricultural production requirements of a growing population, it is essential that we achieve an increased understanding of the basic components and mechanisms shaping growth and productivity of the polyploid wheat plant. Fusarium head blight (FHB) (syn. "scab") poses a serious threat to the quantity and safety of the world's food supply. The resistance locus Fhb1 has provided partial resistance to FHB of wheat for nearly four decades. Map-based cloning of Fhb1 is justified by its significant and consistent effects on reducing disease levels, the importance of FHB in global wheat production and food safety, and because this gene confers partial resistance to this disease and does not appear to behave in a gene-for-gene manner. A bacterial artificial chromosome (BAC) contig spanning the Fhb1 region was developed from the cultivar 'Chinese Spring', sequenced and seven candidate genes were identified in an ~250 kb region. Cosmid clones for each of the seven candidate genes were isolated from a line containing Fhb1 and used for genetic transformation by biolistic bombardment. Transgenic lines were recovered for five candidate genes and evaluated for FHB resistance. All failed to complement the Fhb1 phenotype. Fhb1 is possibly one of the two remaining candidate genes, an unknown regulatory element in this region, or is not present in Chinese Spring.
Traditional views on the effects of polyploidy in allohexaploid wheat have primarily emphasized aspects of coding sequence variation and the enhanced potential to acquire new gene functions through mutation of redundant loci. At the same time, the extent and significance of regulatory variation has been relatively unexplored. Recent investigations have suggested that differential expression of homoeologous transcripts, or subfunctionalization, is common in natural bread wheat. In order to establish a timeline for such regulatory changes and estimate the frequency of non-additive expression of homoeologous transcripts in newly formed T. aestivum, gene expression was characterized in a synthetic T. aestivum line and its T. turgidum and Aegilops tauschii parents by cDNA-SSCP and microarray expression experiments. The cDNA-SSCP analysis of 30 arbitrarily selected homoeologous transcripts revealed that four (~13%) showed differential expression of homoeoalleles in seedling leaf tissue of synthetic T. aestivum. In microarray expression experiments, synthetic T. aestivum gene expression was compared to mid-parent expression level estimates calculated from parental expression levels. Approximately 16% of genes were inferred to display non-additive expression in synthetic T. aestivum. Six homoeologous transcripts classified as non-additively expressed in microarray experiments were characterized by cDNA-SSCP. Expression patterns of these six transcripts suggest that cis-acting regulatory variation is often responsible for non-additive gene expression levels. These results demonstrate that allopolyploidization, per se, results in rapid initiation of differential expression of homoeologous loci and non-additive gene expression in synthetic T. aestivum.
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Genetic characterization of Fusarium head blight resistance in durum wheat / Caractérisation des déterminants génétiques de la résistance à la fusariose chez le blé durPrat, Noémie 28 October 2016 (has links)
La fusariose de l’épi est une maladie fongique qui touche toutes les cultures de céréales à paille à travers le monde entrainant des baisses de rendements et de la qualité des grains. La fusariose pose également un problème pour la sécurité alimentaire lié à la contamination des grains infectés par des mycotoxines. Le développement de variétés résistantes est considéré comme la méthode la plus efficace et la plus durable pour réduire les dommages causés par la maladie et pour limiter la contamination par les mycotoxines. L’amélioration de la résistance à la fusariose chez le blé dur (Triticum durum Desf.) demeure un défi du fait de son extrême sensibilité à la maladie et de la faible variabilité génétique disponible pour ce caractère. L’objectif principal de cette thèse a été d’évaluer l’effet de Fhb1, le QTL majeur de résistance à la fusariose chez le blé tendre (Triticum aestivumL.), au sein de fonds génétiques de blé dur élite. Pour cela, trois populations de cartographie, comprenant chacune environ 100 F7-RIL (lignées pures recombinantes ou « recombinant inbred lines »), ont été développées à partir de croisements entre la lignée expérimentale de blé dur DBC-480, portant une introgression de Fhb1, et les cultivars de blé dur Karur, Durobonus et SZD1029K. Les lignées ont été évaluées au champ, sur trois saisons, pour leur résistance globale à la fusariose après inoculation en spray de Fusarium culmorum. Des notations morphologiques (date de floraison, hauteur des plantes) ont également été réalisées afin d'évaluer leur influence sur l'infestation. Les lignées ont été génotypées à l’aide de marqueurs SSR et de marqueurs GBS (génotypage par séquençage ou « genotyping-by-sequencing ») développés par DArTseq. L’analyse de liaison a permis d’identifier des QTL de résistance sur les bras des chromosomes 2BL, 3BS, 4AL, 4BS, 5AL et 6AS. DBC-480 contribuait à l’allèle de résistance à tous ces loci. Le QTL sur 3BS a été détecté au sein des trois populations centré sur l’intervalle de Fhb1, confirmant, pour la première fois, son introgression dans le blé dur. L’évaluation de la résistance à la propagation après inoculation ponctuelle, réalisé au sein d’une des trois populations, a également permis de valider l’effet de Fhb1 sur la résistance de type 2 chez le blé dur. La hauteur des plantes influe fortement sur la résistance globale à la fusariose et, en particulier, l’allèle de nanisme Rht-B1b est associé à une plus grande sensibilité à la maladie dans les trois populations. Cependant, l’effet négatif de Rht-B1b sur la résistance est largement compensé dans les lignées possédant Fhb1. Des lignées semi-naines avec un meilleur niveau de résistance ont été sélectionnées et favoriseront le développement de cultivars de blé dur résistants à la fusariose. / Fusarium head blight (FHB) is a devastating disease affecting small-grain cereals worldwide causing yield and quality losses. FHB affects food safety due to the contamination of infected grains by mycotoxins. Host plant resistance is considered the most efficient and sustainable approach to contain FHB and mycotoxin contaminations. In durum wheat (Triticum durum Desf.) breeding for FHB resistance remains a challenge due to its extreme susceptibility and to lack of genetic variation available in the primary durum wheat gene pool. The primary goal of this thesis was to evaluate the effect of Fhb1, the major common wheat (Triticum aestivum L.) FHB resistance QTL, in elite durum wheat background. Three F7-RIL (recombinant inbred lines) mapping populations of about 100 lines were developed from crosses between the durum wheat experimental line DBC-480, harboring Fhb1, and the durum wheat cultivars Karur, Durobonus and SZD1029K. The RILs were tested under field conditions by artificial spray inoculation with Fusarium culmorum in three seasons. Morphological traits (flowering date, height) were also recorded to assess their influence on FHB infestation. Genotyping of the lines was performed with SSR and genotyping-by-sequencing (GBS) DArTseq markers. QTL analysis identified genomic regions associated with FHB resistance on chromosome arms 2BL, 3BS, 4AL, 4BS, 5AL and 6AS. DBC-480 contributed the resistant allele at all loci. Fhb1 was detected in all three populations, demonstrating for the first time its successful deployment in durum wheat. The effect of Fhb1 on FHB resistance in durum wheat was further verified by evaluating type 2 resistance in one of the three populations. Plant height had a strong influence in modulating FHB severity. Although the semi-dwarf allele Rht-B1b was associated with increased FHB susceptibility, its negative effect was efficiently counterbalanced in lines carrying Fhb1. Semi-dwarf lines with enhanced levels of resistance were selected and will assist the development of FHB resistant cultivars.
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