A principal concern in crop agriculture is yield, and a key factor for crop growth is the availability of nitrogen. The large amount of nitrogen fertilizer required by plants is a major cost to farmers. Moreover, environmental issues such as groundwater pollution arise from the utilization of nitrogen fertilizers. Therefore, improvement in the nitrogen use efficiency (NUE) of plants is of urgent importance for sustainable and efficient agriculture. Although hybrid varieties have increased crop yields in low N conditions, the molecular mechanism of plant adaptation to N stress is not completely understood. Herein, the study of responses to N limitations in the natural signalling pathways of model plants facilitates the understanding of complex responses in plants to N stress, and this information can be used to further improve NUE. In this research, the transcriptomes of three model plants Arabidopsis, maize, and rice were compared under diverse N growth conditions. An evaluation of the response of the three plants to varying N levels was also conducted. From a statistical point of view, three distinct methods of detecting differential expression were utilized to reduce the likelihood of false positives due to the tens of thousands of genes simultaneously studied. Furthermore, the performance of three statistical approaches was compared during detection of the N-responsive genes. Finally, a clustering analysis (agglomerative hierarchical clustering) was performed on the genes that significantly responded to N levels as identified by a more biologically intuitive method called Rank Products (RP).

Edwardsiella ictaluri is the causative agent of enteric septicemia of catfish (ESC), which is one of the most important diseases impacting the US catfish industry. Though this disease has been very common, progress has been slow to find an economical and practical treatment method. Our long-term goal is to determine the mechanisms of E. ictaluri virulence in ESC. The overall objective of this study was to identify E. ictaluri genes required for host encounter and serum resistance and to determine their roles in pathogenesis. The central hypothesis is that E. ictaluri must differentially regulate its genes to invade fish and evade host defenses, thus, mutation of these differentially expressed genes (DEG) should cause attenuation of E. ictaluri virulence. To test this hypothesis, we first determined the global gene expression patterns of the wild type (wt) E. ictaluri 93-146 and EiAKMut02 mutant during catfish encounter and serum exposure using microarray analysis. Results indicated that in E. ictaluri wt, 377 and 16 DEGs were identified during host encounter and serum exposure, respectively. In EiAKMut02, 82 and 296 DEGs were identified during host encounter and serum experiment. Through functional analysis using Blast2GO, PSORTb, Host Pathogen Interaction Database (HPIDB), and Microbe Virulence Database (MVirDB), 38 DEGs in 9 KEGG pathways have been identified as potential virulence factors. The KEGG pathways represented were 1) bacterial secretion system including T3SS and T6SS, 2) ABC transporters including cystine transport system, iron complex transport system, d-methionine transport system, arginine transport system, thiamine transport system, and molybdate transport system, 3) protein export, 4) flagellar assembly, 5) two-component system, 6) bacterial chemotaxis, 7) ascorbate and aldarate metabolism, 8) phosphotransferase system, and 9) metabolic pathways. In order to understand their role in the E. ictaluri virulence, selected DEGs were inrame deleted by allelic exchange, and their virulence and efficacy were characterized in channel catfish fingerlings. Our results showed that the virulence of E. ictaluri ssaV and yscR mutants was completely attenuated while their efficacies were moderate in catfish fingerlings. These results support that the T3SS and T6SS, ABC transporters, protein export, and flagella seem to be important in E. ictaluri virulence.

A diferenciação morfofisiológica entre rainhas e operárias de Apis mellifera decorre da alimentação recebida durante o desenvolvimento larval, que estimula o aumento da produção de Hormônio Juvenil naslarvas que originarão rainhas. Dentre as diversas diferenças morfológicas entre operárias e rainhas encontramse estruturas especializadas para a coleta de pólen e própolis, localizadas na região da tíbia e do basitarso das pernas posteriores de operárias. A diferenciação das pernas tem início entre o quarto e o quinto estágio do desenvolvimento larval. Utilizandose Microscopia Eletrônica de Varredura o presente trabalho relata a presença das cerdas formando as estruturas castaespecíficas na fase de pupa de olho marrom. A partir de estudos de hibridação de microarrays de cDNA com amostras de RNA de A. mellifera de diversas fases do desenvolvimento larval, foram encontrados 91 genes com ortólogos conhecidos em Drosophila, diferencialmente expressos entre rainhas e operárias no período crítico da diferenciação de castas. Destes, cinco estão relacionados com o desenvolvimento de apêndices: ataxin2 (atx2), cryptocephal (crc), dachshund (dac), grunge (gug) e Retinoic and fat acid Binding Protein (RfaBP). O perfil destes genes, e ainda, ultrabithorax (ubx), distalless(dll) e abdominalA (abdA) (estes porsuassuasfunções durante a diferenciação das pernas de insetos) foram analisados por RTPCR em Tempo Real em pernas posteriores de operárias e rainhas desde o quarto estágio larval até o estágio de pupa de olho branco. Apenas ubx e abdA foram encontrados mais expressos em operárias ao final do desenvolvimento larval e início do desenvolvimento pupal. Estudossimilares dos genes abdA, dac, dll e ubx nossegmentos das pernas de pupas de olho branco indicam a tíbia como domínio de expressão de dac. Imunolocalizações utilizando um anticorpo contra um epitopo conservado entre Ubx e AbdA, FP6.87, em pernas posteriores de prépupas de operárias e rainhasrevelam a presença destas proteínas na tíbia apenas de operárias e diferencialmente localizadas no basitarso de operárias e rainhas. Os dados acima apresentados apontam Ubx, um gene Hox, como pontochave na regulação da formação das estruturas castaespecíficas. / Diphenism in the honey bee, Apis mellifera,resultsfromdifferential feeding of female larvae. Among the morphological differences, the hind legs of workers have structures that is used for carrying pollen and propolis, e.g. the corbicula, while the queens hind legslack thisstructures. The corbicula is an expanded region of the tibia deprived of bristles, which has a single bristle in the middle that seems to have a sensorial function. Using scanning electronic microscopy, we found that the leg structures and bristles of the corbicula are already formed in browneyed pupa. Microarray analysis has demonstrated that five of 240 differentiallyexpressed genesin developing castes are potentially related to the caste differences in leg development (ataxin2, cryptocephal, dachshund, grunge and Retinoic and fat acid Binding Protein). Using qPCR, we analyzed the expression of abdominalA, ataxin2, cryptocephal, grunge, Retinoic and fat acid Binding Protein and ultrabithorax genes during hind leg development. cryptocephal, ataxin2, grunge and Retinoic and fat acid Binding Protein genes, which are involved in imaginal disc elongation and bristle formation and are inhibited by juvenile hormone, were not found to be differentially expressed. However, ultrabithorax and abdominalA are over expressed in workersin the early pupalstage. By using immunohistochemistry, Ubx was localized in the tibia and basitarsus of prepupae of workers and in the basitarsus of pre pupae of queens. The pattern of Ubx expression suggests that this Hox gene is a key player in leg structuresformation and caste differentiation in A.mellifera.

<p>This thesis reports on the development of a novel comprehensive database (Dragon Database of Genes Implicated in Esophageal Cancer, DDEC) as an integrated knowledge database aimed at representing a gateway to esophageal cancer related data. More importantly, it illustrates how the biocurated genes in the database may represent a reliable starting point for divulging transcriptional regulation, diagnostic markers and the biology related to esophageal cancer.</p>

<p>In this thesis the effect of normalization methods on the identification of differentially expressed genes is investigated. A zebrafish microarray dataset called Swirl was used in this thesis work. First the Swirl dataset was extracted and visualized to view if the robust spline and print tip loess normalization methods are appropriate to normalize this dataset. The dataset was then normalized with the two normalization methods and the differentially expressed genes were identified with the LimmaGUI program. The results were then evaluated by investigating which genes overlap after applying different normalization methods and which ones are identified uniquely after applying the different methods. The results showed that after the normalization methods were applied the differentially expressed genes that were identified by the LimmaGUI program did differ to some extent but the difference was not considered to be major. Thus the main conclusion is that the choice of normalization method does not have a major effect on the resulting list of differentially expressed genes.</p>

<p>This thesis reports on the development of a novel comprehensive database (Dragon Database of Genes Implicated in Esophageal Cancer, DDEC) as an integrated knowledge database aimed at representing a gateway to esophageal cancer related data. More importantly, it illustrates how the biocurated genes in the database may represent a reliable starting point for divulging transcriptional regulation, diagnostic markers and the biology related to esophageal cancer.</p>

Philosophiae Doctor - PhD / Esophageal cancer (EC) ranks among the ten most frequent cancers worldwide. Mortality rates associated with EC are very similar to the incidence rates due to the relatively late stage of diagnosis and the poor efficacy of treatment. The aim of this study was to enhance our insights of putative transcriptional circuitry of EC genes, thereby potentially positively impacting our knowledge of therapeutic targets, providing indications as to more appropriate lines of treatment, and additionally allowing for the determination of putative candidate diagnostic markers for the early stage detection of EC. This thesis reports on the development of a novel comprehensive database (Dragon Database of Genes Implicated in Esophageal Cancer, DDEC) as an integrated knowledge database aimed at representing a gateway to esophageal cancer related data. More importantly, it illustrates how the biocurated genes in the database may represent a reliable starting point for divulging transcriptional regulation, diagnostic markers and the biology related to esophageal cancer. DDEC contains known and novel information for 529 differentially expressed EC genes compiled using scientific publications from PubMed and is freely accessible for academic and non-profit users at http://apps.sanbi.ac.za/ddec/. The novel information provided to users of the DDEC is the lists of putative transcription factors that potentially control the 529 manually curated genes. The value of the information accessible through the database was further refined by providing precompiled text-mined and data-mined reports about each of these genes to allow for easy exploration of information about associations of EC-implicated genes with other human genes and proteins, metabolites and enzymes, toxins, chemicals with pharmacological effects, disease concepts and human anatomy. This feature has the capacity to display potential associations that are rarely reported and thus difficult to identify, and it enables the inspection of potentially new ‘association hypotheses’ generated based on the precompiled reports. This study further illustrates how the biocurated esophageal squamous cell carcinoma (ESCC) genes in the database may represent a reliable starting point for exploring beyond current knowledge of the transcriptional circuitry of estrogen related hormone therapy. The genes were used to develop a method that identified 44 combinations of transcription factors (TFs) that characterize the promoter sequence of estrogen responsive genes implicated in ESCC. These significantly over-represented combinations of TFs were then used to increase confidence in the 47 novel putative estrogen response genes that may be related to ESCC too. Coincidently, two of the novel putative estrogen response genes were verified by current (2009), experimental publications. / South Africa

A diferenciação morfofisiológica entre rainhas e operárias de Apis mellifera decorre da alimentação recebida durante o desenvolvimento larval, que estimula o aumento da produção de Hormônio Juvenil naslarvas que originarão rainhas. Dentre as diversas diferenças morfológicas entre operárias e rainhas encontramse estruturas especializadas para a coleta de pólen e própolis, localizadas na região da tíbia e do basitarso das pernas posteriores de operárias. A diferenciação das pernas tem início entre o quarto e o quinto estágio do desenvolvimento larval. Utilizandose Microscopia Eletrônica de Varredura o presente trabalho relata a presença das cerdas formando as estruturas castaespecíficas na fase de pupa de olho marrom. A partir de estudos de hibridação de microarrays de cDNA com amostras de RNA de A. mellifera de diversas fases do desenvolvimento larval, foram encontrados 91 genes com ortólogos conhecidos em Drosophila, diferencialmente expressos entre rainhas e operárias no período crítico da diferenciação de castas. Destes, cinco estão relacionados com o desenvolvimento de apêndices: ataxin2 (atx2), cryptocephal (crc), dachshund (dac), grunge (gug) e Retinoic and fat acid Binding Protein (RfaBP). O perfil destes genes, e ainda, ultrabithorax (ubx), distalless(dll) e abdominalA (abdA) (estes porsuassuasfunções durante a diferenciação das pernas de insetos) foram analisados por RTPCR em Tempo Real em pernas posteriores de operárias e rainhas desde o quarto estágio larval até o estágio de pupa de olho branco. Apenas ubx e abdA foram encontrados mais expressos em operárias ao final do desenvolvimento larval e início do desenvolvimento pupal. Estudossimilares dos genes abdA, dac, dll e ubx nossegmentos das pernas de pupas de olho branco indicam a tíbia como domínio de expressão de dac. Imunolocalizações utilizando um anticorpo contra um epitopo conservado entre Ubx e AbdA, FP6.87, em pernas posteriores de prépupas de operárias e rainhasrevelam a presença destas proteínas na tíbia apenas de operárias e diferencialmente localizadas no basitarso de operárias e rainhas. Os dados acima apresentados apontam Ubx, um gene Hox, como pontochave na regulação da formação das estruturas castaespecíficas. / Diphenism in the honey bee, Apis mellifera,resultsfromdifferential feeding of female larvae. Among the morphological differences, the hind legs of workers have structures that is used for carrying pollen and propolis, e.g. the corbicula, while the queens hind legslack thisstructures. The corbicula is an expanded region of the tibia deprived of bristles, which has a single bristle in the middle that seems to have a sensorial function. Using scanning electronic microscopy, we found that the leg structures and bristles of the corbicula are already formed in browneyed pupa. Microarray analysis has demonstrated that five of 240 differentiallyexpressed genesin developing castes are potentially related to the caste differences in leg development (ataxin2, cryptocephal, dachshund, grunge and Retinoic and fat acid Binding Protein). Using qPCR, we analyzed the expression of abdominalA, ataxin2, cryptocephal, grunge, Retinoic and fat acid Binding Protein and ultrabithorax genes during hind leg development. cryptocephal, ataxin2, grunge and Retinoic and fat acid Binding Protein genes, which are involved in imaginal disc elongation and bristle formation and are inhibited by juvenile hormone, were not found to be differentially expressed. However, ultrabithorax and abdominalA are over expressed in workersin the early pupalstage. By using immunohistochemistry, Ubx was localized in the tibia and basitarsus of prepupae of workers and in the basitarsus of pre pupae of queens. The pattern of Ubx expression suggests that this Hox gene is a key player in leg structuresformation and caste differentiation in A.mellifera.

In this thesis the effect of normalization methods on the identification of differentially expressed genes is investigated. A zebrafish microarray dataset called Swirl was used in this thesis work. First the Swirl dataset was extracted and visualized to view if the robust spline and print tip loess normalization methods are appropriate to normalize this dataset. The dataset was then normalized with the two normalization methods and the differentially expressed genes were identified with the LimmaGUI program. The results were then evaluated by investigating which genes overlap after applying different normalization methods and which ones are identified uniquely after applying the different methods. The results showed that after the normalization methods were applied the differentially expressed genes that were identified by the LimmaGUI program did differ to some extent but the difference was not considered to be major. Thus the main conclusion is that the choice of normalization method does not have a major effect on the resulting list of differentially expressed genes.

No description available.