O conhecimento da diversidade genética e da estrutura populacional das variedades mantidas em bancos de germoplasma é de fundamental importância para sua efetiva utilização em programas de melhoramento. O mapeamento por associação, também conhecido como mapeamento por desequilíbrio de ligação, é um dos principais métodos para relacionar genes e alelos às características de interesse, através da co-segregação de marcadores genéticos polimórficos com os genes envolvidos na variação das características em estudo. O Banco de Germoplasma de Arroz do Departamento de Genética da ESALQ contém 192 acessos japoneses que foram estudados com o objetivo de entender sua diversidade, estruturação genética e determinar a associação genômica de caracteres agronômicos relacionados a produção de grãos. A caracterização molecular foi conduzida através da tecnologia DArT-seq, que gerou dados de marcadores SNPs (single-nucleotide polymorphism) e silico DArTs. Em seguida, após a filtragem, 5.578 SNPs de alta qualidade foram utilizados para calcular as estimativas de diversidade no pacote hierfstat e a estrutura do painel de acessos através da análise discriminante de componentes principais (DAPC), que consiste em determinar existência de cluster em um grupo de genótipos em que não há informação a priori sobre existência de grupos. A diversidade genética nos acessos foi evidenciada pelo valor de heterozigosidade esperada (HS) (0,0279) e a estruturação foi evidenciada pela formação de três subgrupos. O mapeamento associativo foi realizado com o uso do pacote GAPIT, sendo considerados seis caracteres: número de dias para florescimento (NDF), estatura de planta (EP), comprimento da panícula (CP), peso de parcela (PP), massa de mil grãos (MMG) e CICLO, bem como 24.266 marcadores silico DArTs e 1.965 marcadores SNPs. Foram detectadas um total de 113 associações significativas genótipo-fenótipo (P<0,001) quando utilizado marcadores silico DArTs em todas as seis características analisadas e, um total de 21 associações significativas genótipo-fenótipo (P<0,001) quando utilizado marcadores SNPs para apenas quatro das seis características analisadas: EP, CICLO, MMG e PP. Considerando-se os 113 silico DArTs associados significativamente na análise, 90 foram localizados em regiões intergênicas e 23 foram localizados dentro de genes. Enquanto que, dos 21 SNPs significativos, 11 foram localizados em regiões intergênicas e 10 foram localizados dentro de genes. A informação gerada neste estudo foi útil para testar associações ao longo do genoma do arroz. O modelo linear misto (MLM) empregado no mapeamento associativo acredita-se ter conseguido controlar eficientemente os falsos positivos no mapeamento utilizando os marcadores SNPs. As informações geradas neste estudo servem de base para avaliações mais aprofundadas, utilizando o conjunto de marcadores significativos como ponto de partida para determinação dos genes mais importantes para a produtividade em arroz. / The knowledge of the genetic diversity and population structure of varieties maintained in germplasm banks is crucial for their effective use in breeding programs. Association mapping, also known as linkage disequilibrium mapping, is one of the main methods for relating genes and alleles to the characteristics of interest, through the co-segregation of polymorphic genetic markers with the genes involved in the variation of the characteristics under study. The Rice Germplasm Bank of the Department of Genetics of ESALQ contains 192 Japanese accessions that were studied with the purpose of understanding its diversity, genetic structuring and determining the genomic association of agronomic traits related to grain production. The molecular characterization was conducted by DArTseq technology, which generated data of SNPs (single-nucleotide polymorphism) markers and silico DArTs. Then, after filtering, 5,578 high-quality SNPs were used to calculate the diversity estimates in hierfstat package and the accession panel structure through discriminant analysis of principal components (DAPC), which consists of determining the cluster existence in a group of genotypes where there is no a priori information about the existence of groups. The genetic diversity in the accessions was evidenced by the expected heterozygosity value (HS) (0.0279) and the population structure was evidenced by the formation of three clusters. The association mapping was performed using the GAPIT package, considering six characters: number of days for flowering (NDF), plant height (EP), panicle length (CP), plot weight (PP), mass of thousand grains (MMG) and CYCLE, as well as 24.266 silico DArTs markers and 1.965 SNPs markers. We detected a total of 113 significant associations genotype-phenotype (P <0.001) when used silico DArTs markers in all six analyzed characteristics and a total of 21 significant associations genotype-phenotype (P<0.001) when used SNPs markers for only four of the six analyzed characteristics: EP, CYCLE, MMG and PP. Considering the 113 silico DArTs significantly associated in the analysis, 90 were located in intergenic regions and 23 were localized within genes. While of the 21 significant SNPs, 11 were located in intergenic regions and 10 were located within genes. The information generated in this study was useful for testing associations throughout the rice genome. The mixed linear model (MLM) used in association mapping is believed to have been able to efficiently control false positives in the mapping using the SNPs markers. The information generated in this study serves as a basis for further evaluation using the set of significant markers as a starting point for determining the most important genes for rice yield.

Made available in DSpace on 2014-06-11T19:30:59Z (GMT). No. of bitstreams: 0 Previous issue date: 2010-12-06Bitstream added on 2014-06-13T18:41:21Z : No. of bitstreams: 1 almeida_rba_dr_sjc.pdf: 7133488 bytes, checksum: a3104ef679cdc446d58a95b507c26864 (MD5) / Na procura de meios preventivos e curativos para doença periodontal e cárie dentária, plantas medicinais com finalidade fungicida, bactericida e antiinflamatória vêm sendo investigadas. O objetivo desse trabalho foi o de avaliar a atividade antimicrobiana utilizando-se óleos essenciais de Cymbopogon citratus (D.C.) Stapf, Curcuma zedoaria Roscoe e Tagetes minuta L. sobre cepas de Staphylococcus spp., Streptococcus mutans e Candida spp. em crescimento planctônico e em biofilme. Para estudo dos microrganismos em crescimento planctônico, foram determinadas a Concentração Inibitória Mínima e a Concentração Fungicida Mínima de nove cepas clínicas e uma cepa padrão para cada espécie: C. albicans, C. tropicalis, C. glabrata, S. aureus, S. epidermidis e S. mutans. Para avaliação dos efeitos dos óleos essenciais em biofilme, foram utilizadas cepas padrão de Candida albicans (ATCC 18804), Staphylococcus aureus (ATCC 6538) e Streptococcus mutans (ATCC 35688) isolados e em associações em corpos-de-prova durante cinco dias. A seguir, os corpos-de-prova em resina acrílica foram lavados e colocados em contato com óleos essenciais durante 5 min. O número de unidades formadoras de colônias obtidas em cada biofilme (UFC/mL) foram submetidos à análise estatística descritiva e teste t-Student utilizando-se o software Minitab considerando-se o nível de significância p ≤ 0,05. Também foi realizada microscopia eletrônica de varredura (MEV) nos corpos-de-prova em resina acrílica com biofilme com tratamento e controle. Foram observadas reduções significativas do número de unidades formadoras de colônias (UFC/mL) tanto no biofilme como em associação. As maiores reduções ocorreram no tratamento do óleo essencial de C. citratus, seguidas pelo óleo de T. minuta e C. zedoaria. Os óleos essenciais de Cymbopogon citratus, Tagetes minuta e Curcuma zedoaria apresentaram... / In the search for preventive and curative means for periodontal disease and tooth decay, herbal-purpose fungicide, antibacterial and anti-inflammatory have been investigated. The aim of this study was to evaluate the antimicrobial activity using the essential oils of Cymbopogon citratus (D.C.) Stapf, Curcuma zedoaria Roscoe and Tagetes minuta L. on strains of Staphylococcus spp., Streptococcus mutans and Candida spp. in planktonic and biofilm growth. For the study of microorganisms in planktonic growth, were determined Minimum Inhibitory Concentration and Minimum Microbicidal Concentration of nine clinical strains and one ATCC for each species: C. albicans, C. tropicalis, C. glabrata, S. aureus, S. epidermidis and S. mutans. To assess the effects of essential oils in biofilm, was used strains of Candida albicans (ATCC 18804), Staphylococcus aureus (ATCC 6538) and Streptococcus mutans (ATCC 35688) alone and in associations in acrylic resin discs immersed in sterile brain heart infusion broth (BHI) containing 5% sucrose, inoculated with microbial suspension and incubated for 5 days. On the fifth day, the discs were washed in sterile saline solution in order to remove loosely bound material. Next, the acrylic resin discs were washed and placed in contact with essential oils for 5 min. The number of CFU obtained in each biofilm (CFU/mL) were subjected to descriptive statistical analysis using Minitab software considering with significance level p ≤ 0.05. It was also performed scanning electron microscopy (SEM) acrylic resin disc with biofilm with treatment and control. It was identified significant reductions in the number of colony forming units (CFU/mL) in both planktonic and biofilm associated. The largest reductions occurred in the treatment of essential oil of C. citratus, followed by oil of T. minuta and C. zedoaria. We conclude that the essential oils of Cymbopogon citratus... (Complete abstract click electronic access below)

<p>Human disturbance has led to excessive deforestation and to a very limited forest cover in the Afromontane zone of Ethiopia, which forms a large part of the country. Thus urgent conservation measures are required to ameliorate the situation. Understanding the natural regeneration processes and the dynamics of plant populations of tree and shrub species has a practical application in the restoration of these habitats. </p><p>The present study focuses on the population dynamics and regeneration of an early successional shrub <i>Dodonaea angustifolia</i> and a late successional tree <i>Olea europaea</i> ssp. <i>cuspidata </i>in southern Wello, Ethiopia. Population structure and dynamics, regeneration and seed banks in dry Afromontane habitats were considered. </p><p>For both species, three population structure patterns were identified: 1) high density, reversed J-shape structure with many seedlings and few large individuals, 2) lower density, unimodal structure with higher proportions of plants of intermidiate size, 3) high density with higher proportions of large plants , in some cases bimodal with small and large individuals. Vegetation type and protection time were found to have a significant effect on the population structure of both species. <i>Dodonaea</i> can establish itself on degraded land, once the disturbance has ceased</p><p>Projection matrix analysis on observations from permanent plots in <i>Dodonaea</i> populations in protected and unprotected sites resulted in one declining population, and one increasing in the protected site and declining populations at the unprotected site. The overall projected growth rate in <i>Dodonaea</i> calculated from a pooled matrix indicated positive population growth. The factors influencing the population growth, recruitment and survival are discussed.</p><p>The persistence of <i>Olea </i>populations seems to depend on the more stable environmental conditions in later successional stages of forest vegetation. There are possibilities of natural regeneration of <i>Olea</i> if regenerating individuals still occur in the area. Rainfall seasonality is a dominant factor in regulating establishment, recruitment, survival and growth, particularly during the seedling stage. Moreover, shade and herbivory are factors that need consideration. Since <i>Olea</i> grows better under shade than in the open sun, successful regeneration for this species relies on shade from other plants and on protection from grazing, at least during the seedling stage.</p><p>Most of the species that germinated from the seed banks were herbs and grasses with very few shrub and tree species. There was low correspondence between species composition of the seed banks and that of the standing vegetation. </p><p>Spatial and temporal variation in demographic parameters among populations of <i>Dodonaea</i> and <i>Olea </i>can be attributed to human and environmental influence. Under protection, both <i>Dodonaea</i> and <i>Olea</i> seem to have a possibility to regenerate naturally. Further research should consider factors mentioned in detailed investigations of other dominant Afromontane forest species.</p>

Human disturbance has led to excessive deforestation and to a very limited forest cover in the Afromontane zone of Ethiopia, which forms a large part of the country. Thus urgent conservation measures are required to ameliorate the situation. Understanding the natural regeneration processes and the dynamics of plant populations of tree and shrub species has a practical application in the restoration of these habitats. The present study focuses on the population dynamics and regeneration of an early successional shrub Dodonaea angustifolia and a late successional tree Olea europaea ssp. cuspidata in southern Wello, Ethiopia. Population structure and dynamics, regeneration and seed banks in dry Afromontane habitats were considered. For both species, three population structure patterns were identified: 1) high density, reversed J-shape structure with many seedlings and few large individuals, 2) lower density, unimodal structure with higher proportions of plants of intermidiate size, 3) high density with higher proportions of large plants , in some cases bimodal with small and large individuals. Vegetation type and protection time were found to have a significant effect on the population structure of both species. Dodonaea can establish itself on degraded land, once the disturbance has ceased Projection matrix analysis on observations from permanent plots in Dodonaea populations in protected and unprotected sites resulted in one declining population, and one increasing in the protected site and declining populations at the unprotected site. The overall projected growth rate in Dodonaea calculated from a pooled matrix indicated positive population growth. The factors influencing the population growth, recruitment and survival are discussed. The persistence of Olea populations seems to depend on the more stable environmental conditions in later successional stages of forest vegetation. There are possibilities of natural regeneration of Olea if regenerating individuals still occur in the area. Rainfall seasonality is a dominant factor in regulating establishment, recruitment, survival and growth, particularly during the seedling stage. Moreover, shade and herbivory are factors that need consideration. Since Olea grows better under shade than in the open sun, successful regeneration for this species relies on shade from other plants and on protection from grazing, at least during the seedling stage. Most of the species that germinated from the seed banks were herbs and grasses with very few shrub and tree species. There was low correspondence between species composition of the seed banks and that of the standing vegetation. Spatial and temporal variation in demographic parameters among populations of Dodonaea and Olea can be attributed to human and environmental influence. Under protection, both Dodonaea and Olea seem to have a possibility to regenerate naturally. Further research should consider factors mentioned in detailed investigations of other dominant Afromontane forest species.

The contamination of seafood with methylmercury (MeHg) is a global health issue, as MeHg is a well known neurotoxin. Since dietary nutrients may interact with MeHg toxicity, and oxidative stress is one of the primary mechanisms underlying MeHg neurotoxicity, we characterized dietary antioxidant-MeHg interactions. Firstly, we used an ethnobotanical study to confirm the antioxidant activity of Northern Labrador Tea, Rhododendron tomentosum ssp. subarcticum (Tea), for the Canadian Inuit, a population with elevated MeHg exposure. Secondly, we determined the ability of Tea to ameliorate MeHg-induced toxicity in a rat perinatal exposure study. MeHg exposure (2 mg/KgBW/d) was associated with perturbed development and behaviour, elevated brain N-methyl-D-aspartate receptors, and serum lipid peroxidation. Surprisingly, Tea co-exposure (100 mg/KgBW/d) modulated MeHg’s effects on brain NMDA-R levels and lipid peroxidation, but also increased mercury serum concentrations. Thirdly, using a toxicogenomics approach we determined that MeHg exposure caused the down-regulation of Nr4a2 and its protein product Nurr1. These novel MeHg targets are implicated in developmental learning functions and were corrected with MeHg + Tea co-exposure. Lastly, we conducted a risk assessment survey and cross-sectional dietary epidemiology study in Costa Rica to further investigate dietary nutrient-MeHg interactions. Costa Rica is a Central American country with multiple sources of Hg and a high per capital fish consumption. Here, 5 of the 14 populations we studied exceeded the recommended MeHg provisional tolerable daily intake (pTDI) of 0.2 µg/KgBW/d. In Heredia the pTDI was exceeded by 34% of woman participants, primarily associated with canned tuna consumption. Interestingly, we detected that Hg body burden was significantly reduced by the consumption of antioxidant-rich dietary items. Considering our collective results, we hypothesized that MeHg toxicokinetics may be altered by dietary nutrients at the site of intestinal absorption from the disruption of gut flora, or at the site of cellular demethylation in tissues from the improvement of cellular redox state. The interaction of dietary nutrients on MeHg outcomes has a large impact on risk assessment and may provide a public health approach for managing the risk associated with MeHg exposure without reducing local fish consumption.

Against the background of past, recent and future climate change, the present thesis addresses elevational shifts of alpine treelines in the Swedish Scandes. By definition, treeline refers to the elevation (m a.s.l.) at a specific site of the upper trees of a specific tree species, at least 2 m tall. Based on historical records, the first part of the thesis reports and analyzes the magnitude of treeline displacements for the main trees species (Betula pubescens ssp. czerepanovii, Picea abies and Pinus sylvestris) since the early 20th century. The study covered a large and heterogeneous region and more than 100 sites. Concurrent with temperature rise by c. 1.4 °C over the past century, maximum treeline advances of all species amount to about 200 m. That is virtually what should be predicted from the recorded temperature change over the same period of time. Thus, it appears that under ideal conditions, treelines respond in close equilibrium with air temperature evolution. However, over most parts of the landscape, conditions are not that ideal and treeline upshifts have therefore been much smaller. The main reason for that discrepancy was found to be topoclimatic constraints, i.e. the combined action of geomorphology, wind, snow distribution, soil depth, etc., which over large parts of the alpine landscape preclude treelines to reach their potential thermal limit. Recorded treeline advance by maximum 200 m or so over the past century emerges as a truly anomalous event in late Holocene vegetation history. The second part of the thesis is focused more on long-term changes of treelines and one specific and prevalent mechanism of treeline change. The first part of the thesis revealed that for Picea and Betula, treeline shift was accomplished largely by phenotypic transformation of old-established stunted and prostrate individuals (krummholz) growing high above the treeline. In obvious response to climate warming over the past century, such individuals have transformed into erect tree form, whereby the treeline (as defined here) has risen. As a means for deeper understanding of this mode of positional treeline change, extant clonal spruces, growing around the treeline, were radiocarbon dated from megafossil remains preserved in the soil underneath their canopies. It turned out that Picea abies in particular may attain almost eternal life due to its capability for vegetative reproduction and phenotypic plasticity. Some living clones were in fact inferred to have existed already 9500 years ago, and have thus persisted at the same spot throughout almost the entire Holocene. This contrasts with other tree species, which have left no living relicts from the early Holocene, when they actually grew equally high as the spruce. Thereafter they retracted by more than 300 m in elevation supporting that also on that temporal scale, treelines are highly responsive to climate change. The early appearance of Picea in the Scandes, suggests that Picea “hibernated” the last glacial phase much closer to Scandinavia than earlier thought. It has also immigrated to northern Sweden much earlier than the old-established wisdom. The experiences gained in this thesis should constitute essential components of any model striving to the project landscape ecological consequences of possible future climate shifts.

The contamination of seafood with methylmercury (MeHg) is a global health issue, as MeHg is a well known neurotoxin. Since dietary nutrients may interact with MeHg toxicity, and oxidative stress is one of the primary mechanisms underlying MeHg neurotoxicity, we characterized dietary antioxidant-MeHg interactions. Firstly, we used an ethnobotanical study to confirm the antioxidant activity of Northern Labrador Tea, Rhododendron tomentosum ssp. subarcticum (Tea), for the Canadian Inuit, a population with elevated MeHg exposure. Secondly, we determined the ability of Tea to ameliorate MeHg-induced toxicity in a rat perinatal exposure study. MeHg exposure (2 mg/KgBW/d) was associated with perturbed development and behaviour, elevated brain N-methyl-D-aspartate receptors, and serum lipid peroxidation. Surprisingly, Tea co-exposure (100 mg/KgBW/d) modulated MeHg’s effects on brain NMDA-R levels and lipid peroxidation, but also increased mercury serum concentrations. Thirdly, using a toxicogenomics approach we determined that MeHg exposure caused the down-regulation of Nr4a2 and its protein product Nurr1. These novel MeHg targets are implicated in developmental learning functions and were corrected with MeHg + Tea co-exposure. Lastly, we conducted a risk assessment survey and cross-sectional dietary epidemiology study in Costa Rica to further investigate dietary nutrient-MeHg interactions. Costa Rica is a Central American country with multiple sources of Hg and a high per capital fish consumption. Here, 5 of the 14 populations we studied exceeded the recommended MeHg provisional tolerable daily intake (pTDI) of 0.2 µg/KgBW/d. In Heredia the pTDI was exceeded by 34% of woman participants, primarily associated with canned tuna consumption. Interestingly, we detected that Hg body burden was significantly reduced by the consumption of antioxidant-rich dietary items. Considering our collective results, we hypothesized that MeHg toxicokinetics may be altered by dietary nutrients at the site of intestinal absorption from the disruption of gut flora, or at the site of cellular demethylation in tissues from the improvement of cellular redox state. The interaction of dietary nutrients on MeHg outcomes has a large impact on risk assessment and may provide a public health approach for managing the risk associated with MeHg exposure without reducing local fish consumption.

The contamination of seafood with methylmercury (MeHg) is a global health issue, as MeHg is a well known neurotoxin. Since dietary nutrients may interact with MeHg toxicity, and oxidative stress is one of the primary mechanisms underlying MeHg neurotoxicity, we characterized dietary antioxidant-MeHg interactions. Firstly, we used an ethnobotanical study to confirm the antioxidant activity of Northern Labrador Tea, Rhododendron tomentosum ssp. subarcticum (Tea), for the Canadian Inuit, a population with elevated MeHg exposure. Secondly, we determined the ability of Tea to ameliorate MeHg-induced toxicity in a rat perinatal exposure study. MeHg exposure (2 mg/KgBW/d) was associated with perturbed development and behaviour, elevated brain N-methyl-D-aspartate receptors, and serum lipid peroxidation. Surprisingly, Tea co-exposure (100 mg/KgBW/d) modulated MeHg’s effects on brain NMDA-R levels and lipid peroxidation, but also increased mercury serum concentrations. Thirdly, using a toxicogenomics approach we determined that MeHg exposure caused the down-regulation of Nr4a2 and its protein product Nurr1. These novel MeHg targets are implicated in developmental learning functions and were corrected with MeHg + Tea co-exposure. Lastly, we conducted a risk assessment survey and cross-sectional dietary epidemiology study in Costa Rica to further investigate dietary nutrient-MeHg interactions. Costa Rica is a Central American country with multiple sources of Hg and a high per capital fish consumption. Here, 5 of the 14 populations we studied exceeded the recommended MeHg provisional tolerable daily intake (pTDI) of 0.2 µg/KgBW/d. In Heredia the pTDI was exceeded by 34% of woman participants, primarily associated with canned tuna consumption. Interestingly, we detected that Hg body burden was significantly reduced by the consumption of antioxidant-rich dietary items. Considering our collective results, we hypothesized that MeHg toxicokinetics may be altered by dietary nutrients at the site of intestinal absorption from the disruption of gut flora, or at the site of cellular demethylation in tissues from the improvement of cellular redox state. The interaction of dietary nutrients on MeHg outcomes has a large impact on risk assessment and may provide a public health approach for managing the risk associated with MeHg exposure without reducing local fish consumption.

Banana is the most important fruit crop in the world but ironically one of the crops least studied. This fruit constitutes a major staple food for millions of people in developing countries and also it is considered the highest selling fruit in the world market making this crop a very important export commodity for the producing countries. At the present time, one of the most significant constraints of banana production that causes significant economical losses are fungal diseases. Among these, Panama disease, also known as Fusarium wilt has been the most catastrophic. Panama disease is caused by the soil-borne fungus Fusarium oxysporum formae specialis (f.sp) cubense (FOC), which infects susceptible bananas through the roots causing a lethal vascular wilt. To date, the race 4 of this pathogen represents the most serious threat to banana production worldwide since most of the commercial cultivars are highly susceptible to this pathogen. Introduction of FOC resistance into commercial cultivars by conventional breeding has been difficult because edible bananas are sterile polyploids without seeds. Genetic transformation of banana, which has already been established in various laboratories around the world has the potential to solve this problem by transferring a FOC race 4 resistance gene into susceptible banana cultivars (eg. Cavendish cultivars). However, a FOC resistant (R) gene has not been isolated. Genes that confer resistance to Fusarium oxysporum have been isolated from tomato and melon using a map-based positional cloning approach. The tomato I2 and melon Fom-2 genes belong to the non-Toll/interleukin like receptors (TIR) subclass of nucleotide-binding site and leucine-rich repeat (NBS-LRR) R genes. These genes confer resistance only to certain races of F. oxysporum in their corresponding plant families limiting their use in other plant families. The fact that these two Fusarium resistance genes share the same basic non-TIR-NBS-LRR structure suggests a similar Fusarium resistance mechanism is shared between the families Solanaceae and Cucurbitaceae. This observation opens the possibility to find similar Fusarium resistance genes in other plant families including the Musaceae. A remarkable discovery of a population of the wild banana Musa acuminata subspecies (ssp.) malaccensis segregating for FOC race 4 resistance was made by Dr. Ivan Buddenhagen (University of California, Davis) in Southeast Asia. Research carried out at Queensland Department of Primary Industries (Australia) using this plant material has demonstrated that a single dominant gene is involved in FOC race 4 resistance (Dr. Mike Smith, unpublished results). Tissue-culture plantlets of this FOC race 4 segregating population were kindly provided to the Plant Biotechnology Program (Queensland University of Technology) by Dr. Mike Smith to be used in our research. This population holds the potential to assist in the isolation of a FOC race 4 resistance gene and other potential Fusarium resistance genes. The overall aims of this research were to isolate and characterise resistance gene candidates of the NBS-type from M. acuminata ssp. malaccensis and to identify and characterise potential Fusarium resistance genes using a combination of bioinformatics and gene expression analysis. Chapter 4 describes the isolation by degenerate PCR of five different classes of NBS sequences from banana (Musa acuminata ssp malaccensis) designated as resistance gene candidates (RGCs). Deduced amino acid sequences of the RGCs revealed the typical motifs present in the majority of known plant NBS-LRR resistance genes. Structural and phylogenetic analyses showed that the banana RGCs are related to non-TIR subclass of NBS sequences. The copy number of each class was estimated by Southern hybridisation and each RGC was found to be in low copy number. The expression of the RGCs was assessed by RT-PCR in leaf and root tissues of plants resistant or susceptible to Fusarium oxysporum f. sp. cubense (FOC) race 4. Four classes showed a constitutive expression profile whereas no expression was detected for one class in either tissue. Interestingly, a transcriptional polymorphism was found for RGC2 whose expression correlated with resistance to FOC race 4 suggesting a possible role of this gene in resistance to this devastating FOC race. Moreover, RGC2 along with RGC5 showed significant sequence similarity to the Fusarium resistance gene I2 from tomato and were chosen for further characterisation. The NBS sequences isolated in this study represent a valuable source of information that could be used to assist the cloning of functional R genes in banana. Chapter 5 describes the isolation and characterisation of the full open reading frame (ORF) of RGC2 and RGC5 cDNAs. The ORFs of these two banana RGCs were predicted to encode proteins that showed the typical structure of non-TIR-NBS-LRR resistance proteins. Homology searches using the entire ORF of RGC2 and RGC5 revealed significant sequence similarity to the Fusarium resistance gene I2 from tomato. Interestingly, the phylogenetic analysis showed that RGC2 and RGC5 were grouped within the same phylogenetic clade, along with the Fusarium resistance genes l2 and Fom-2. These findings suggest that the banana RGC2 and RGC5 are potential resistance gene candidates that could be associated with Fusarium resistance. The case of RGC2 is more remarkable because its expression was correlated to FOC race 4 resistance (Chapter 4). As a first step to test whether RGC2 has a role in FOC race 4 resistance, different expression constructs were made with the ORF of this sequence. One of the constructs contains a RGC2 putative promoter region that was successfully cloned in this work. These constructs will be used to transform susceptible banana plants that can then be challenged with FOC race 4 to assess whether resistance has been acquired by genetic complementation. The results of this thesis provide interesting insights about the structure, expression and phylogeny of two potential Fusarium resistance genes in banana, and provide a rational starting point for their functional characterisation. The information generated in this thesis may lead to the identification of a Fusarium resistance gene in banana in further studies and may also assist the cloning of Fusarium resistance genes in other plant species.

Production errors made by second language (L2) learners of English have been attributed to markedness, L1 transfer or input frequency (cf. Major, 2001; Edwards & Zampini, 2008; Baptista, Rauber, & Watkins, 2009). This thesis examines the production of 17 English initial consonant clusters (e.g., /pr/ in “pray”) in a markedness relationship, whereby clusters with greater sonority distance between the first and second consonants are unmarked and clusters with smaller sonority distance between the first and second consonants are marked, by two groups of Saudi Arabian L2 English learners. It also explores the effect of input frequency and L1 transfer. Participants were asked to read 60 sentences and their reading was recorded for acoustic analysis. Analysis showed that “prothesis” was always used to simplify the clusters, and that, the duration of the prothetic vowel tended to get longer when clusters become more marked. Intermediate participants had greater degree of difficulty in producing the clusters and tended to insert a longer prothetic vowel in general. Markedness explained the performance on #sC clusters; however, performance on non #sC clusters was best explained by L1 transfer. Results further indicated that input frequency was irrelevant to this study.