Made available in DSpace on 2014-06-11T19:30:55Z (GMT). No. of bitstreams: 0 Previous issue date: 2011-02-24Bitstream added on 2014-06-13T18:41:10Z : No. of bitstreams: 1 rodovalho_cm_dr_rcla.pdf: 1352309 bytes, checksum: b91a746d690f5903c39601146a591fab (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Formigas cortadeiras do gênero Atta, popularmente conhecidas como saúvas, são as mais derivadas dentro da tribo Attini. Apresentam grande importância ecológica, porém, pelo hábito de cortarem folhas para manutenção do fungo simbionte e pelo enorme tamanho das colônias, causam muitos prejuízos às lavouras, pastagens e plantações, sendo consideradas pragas agrícolas. Atta laevigata Smith, 1858 apresenta vasta distribuição pelo Brasil e é responsável pela herbivoria de inúmeras plantas dicotiledôneas, gramíneas e espécies nativas de diferentes biomas. O presente trabalho teve como objetivos a caracterização parcial do transcriptoma e do genoma mitocondrial de A. laevigata. Foram caracterizadas 2006 sequências únicas do transcriptoma, a partir de uma biblioteca de cDNA preparada com indivíduos inteiros da formiga. Entre essas sequências, 16 provavelmente representam genes com grande número de transcritos. Esses 16 genes estão relacionados a três funções celulares: (i) conservação de energia através de reações redox na mitocôndria; (ii) estrutural, pelo citoesqueleto e músculos; (iii) regulação da expressão gênica e metabolismo. Considerando o estilo de vida e processos biológicos chaves para essas formigas, 146 sequências foram identificadas com base na sua utilização para o controle de cortadeiras pragas. A partir de dados da biblioteca de cDNA e procedimentos envolvendo primer walking, o genoma mitocondrial de A. laevigata foi parcialmente caracterizado, apresentandose com 17920 pb, maior, portanto, do que outros já descritos em Hymenoptera, mesmo considerando-se a impossibilidade de determinação da sequência de uma pequena porção do mtDNA, envolvendo a região controle, uma parte do 12S e os tRNAs S1, V e M. Como já descrito para outros mitogenomas, o de A. laevigata apresentou alto conteúdo AT, os mesmos 13 genes codificadores... / Leafcutter ants from Atta genus, popularly known as “saúvas”, are the most derived of the tribe Attini. They have major ecological importance, but, because of their habit of cutting leaves for the maintenance of the symbiotic fungus and the huge colony size, they impose severe economic damages to plantations, pastures, and agriculture, being considered as agriculture pests. Atta laevigata shows wide distribution in Brazil and it is responsible for the herbivory of many dicots, grass, and native species from different biomes. The present work aimed to characterize the transcriptome and the mitochondrial genome of A. laevigata. 2,006 unique sequences of the transcriptome were characterized from a cDNA library constructed with whole individuals. Among those sequences, 16 are likely from genes with high number of transcripts. Those 16 genes are related with three cellular functions: (i) energy conservation through redox reactions in mitochondria; (ii) cytoskeleton and muscle structuring; (iii) regulation of gene expression and metabolism. Based on lifestyle and key biological processes of these ants, 146 sequences were identified with potential use for controlling pest leafcutters. Using data from cDNA library and primer walking proceedings, the mitochondrial genome of A. laevigata was partially characterized with 17,920 bp, being larger than the others already described for Hymenoptera. A small part of the mtDNA was not sequenced, including the control region, a portion of 12S and tRNAs S1, V, and M. As described before for other mitogenomes, A. laevigata mtDNA displayed high AT contain, the same 13 proteincoding genes and the two ribosomal subunits with length and location according to the hypothetic ancestral mitogenome. Rearrangements were found for the tRNAs, but the most remarkable difference were the high number and longer length of intergenic regions presented in the mtDNA... (Complete abstract click electronic access below)

No description available.

Made available in DSpace on 2014-06-11T19:26:09Z (GMT). No. of bitstreams: 0 Previous issue date: 2007-07-20Bitstream added on 2014-06-13T18:54:09Z : No. of bitstreams: 1 jovino_dfr_me_jabo.pdf: 637580 bytes, checksum: dc449a2a52a9354da867effed1512963 (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Sob condições de estresse hídrico, a cana-de-açúcar pode sofrer mudanças fisiológicas e bioquímicas, tais como diminuição nas atividades fotoquímicas, redução da fixação de CO2 e acúmulo de osmólitos e osmoprotetores. O objetivo deste trabalho foi identificar, através da técnica de macroarranjo de cDNA, o perfil de expressão de genes promotores das diferentes vias metabólicas em palmitos da variedade de cana-de-açúcar SP80-3280 submetidas ao estresse hídrico nos dias 5, 9, 13 e 17 após o início da condição de supressão de água, sendo considerado o dia 1 como controle. Os resultados do macroarranjo mostraram que as proteínas mais expressas sob déficit hídrico pertencem a quatro categorias das quais as ESTs mais importantes foram selecionadas. As quatro categorias descritas abaixo estão discutidas neste trabalho. As ESTs da via do metabolismo de açúcar e amido (invertase de parede celular (INV), sacarose fosfato sintase (SFS), sacarose fosfato fosfatase (SFF), trealose fosfato sintase (TFS), trealose fosfato sintase/fosfatase (TFS/F) e hexoquinase (HXQ)) pertencem a categoria de bioenergética. Colina monooxigenase (CMO) e betaína aldeído desidrogenase (BADH) as quais pertencem a via do metabolismo de glicina betaína, foram selecionadas a partir da categoria do metabolismo secundário. A terceira categoria compreende as ESTs do metabolismo de aminoácido (D-pirrolina-5- carboxilato sintase (P5CS), ornithina -aminotransferase (OAT) e prolil 4-hidroxilase (PH)) as quais pertencem a via da biossíntese de prolina, e a quarta categoria, resposta ao estresse, compreende as ESTs aleno oxido sintase (AOS), aleno oxido ciclase (AOC) e lipoxigenase (LOX), pertencentes a via de biossíntese do jasmonato. / Under water deficit, sugarcane undergoes physiological and biochemical alterations such as photochemical activity reduction, CO2 fixation reduction and the accumulation of osmolytes and osmoprotectants. This work was undertaken to identify the gene expression profile in sugarcane (var. SP80-3280) under water deficit through the cDNA macroarray technique. Leafroll tissues were collected from plants subjected to 5, 9, 13 and 17 days of water restriction, and day 1 was used as control. Macroarray results showed that most proteins expressed under water restriction belong to four categories, from which the most important ESTs were selected. The four categories described below are discussed in this work. Sugar and starch metabolism pathway ESTs (cell wall invertase (CWI), sucrose phosphate synthase (SPS), sucrose phosphate phosphatase (SPP), trehalose phosphate synthase (TPS), trehalose phosphate synthase phosphatase (TSP/P) and hexoquinase (HXQ) were selected from the bioenergetics category. Choline monooxygenase (CMO) and betaine aldehyde dehydrogenase (BADH), which belong to the glycine betaine metabolism pathway, were selected from the secondary metabolism category. The third category comprised the amino acid metabolism ESTs D- pyrroline-5-carboxylate synthase (P5CS), Ornithine - aminotransferase (OAT) and Prolyl 4-hydroxylase (PH), which belong to the proline biosynthesis pathway, and the fourth category, stress response, comprised the ESTs for Allene oxide synthase (AOS), Allene oxide cyclase (AOC) and Lipoxygenase (LOX), which belong to the jasmonate biosynthesis pathway.

RNAs não codificadores longos (lncRNAs) compõem uma fração significativa do transcriptoma. Alterações na expressão de lncRNAs já foram observadas em vários cânceres humanos, mas ainda não foram exploradas no adenocarcinoma pancreático ductal (PDAC), uma doença devastadora e agressiva para a qual faltam métodos para diagnóstico precoce e tratamentos efetivos. Utilizando uma plataforma de microarranjo de cDNA com sondas para 984 lncRNAs e 2371 mRNAs, o presente estudo identificou conjuntos de lncRNAs expressos em 38 amostras clínicas pancreáticas. O enriquecimento de (i) elementos regulatórios associados às regiões promotoras (H3K4me3); (ii) possíveis inícios de transcrição (CAGE-tags); (iii) presença de elementos conservados sugere que ao menos uma fração desses RNAs seja originada a partir de unidades transcricionais independentes, reguladas e possivelmente funcionais. Foram identificadas assinaturas de expressão gênica compostas por mRNA e lncRNAs associadas ao tumor primário e à metástase pancreática. A assinatura gIenica associada à metástase apresentou enriquecimento RNAs intrônicos de loci gênicos associados à via MAPK quinase. O aumento de expressão dos transcritos intrônicos dos loci PPP3CB, MAP3K14 e DAPK1 foi confirmado por qPCR em metástases. Em conjunto, este trabalho aponta para a importância de lncRNAs intrônicos no PDAC e para a necessidade de estudos mais aprofundados para uma melhor compreensão do papel dessa classe de transcritos na biologia da doença. / Long noncoding RNAs (lncRNAs) compose a significant fraction of transcriptome. Altered expression of lncRNAs has been observed in diverse human cancers, but has not being investigated in pancreatic ductal adenocarcinoma (PDAC), a devastating and aggressive disease that lack early diagnosis methods and effective treatments. Using a cDNA microarray platform with probes interrogating 984 lncRNAs and 2371 mRNA, the present study identified subsets of lncRNAs expressed in 38 pancreatic clinical samples. Enrichment of (i) regulatory elements associated to promoter region (H3K4me3); (ii) putative transcription start site (CAGEtags) and (iii) conserved elements, suggest that at least a fraction of these RNAs could be independent transcriptional unit, regulated, an possibly functional. Gene expression signatures comprised of mRNAs and lncRNAs and associated to primary or metastatic tumors were found. A gene signature associated to metastasis was enriched in intronic ncRNAs mapping to gene loci associated to the MAPK pathway. Over expression of intronic RNAs from PPP3CB, MAP3K14 and DAPK1 was confirmed by qPCR in metastatic samples. Taken together, this study points to the importance of intronic lncRNAs in PDAC and for the need to study this class of ncRNAs in greater detail to better understand its role in the biology of PDAC.

Made available in DSpace on 2014-08-20T13:44:42Z (GMT). No. of bitstreams: 1 dissertacao_liliane_mertz.pdf: 5055852 bytes, checksum: b6922bc49af5dfa5f81f10ad3f997443 (MD5) Previous issue date: 2007-03-22 / Some works have reported the existence of contrasting soybean genotypes for physiological quality of seeds. There are such differences because of the presence of seeds with total or partial impermeability for water absorption into the coat that make them less susceptible to mechanical damages, weather adversities, deterioration by humidity and pathogen occurrence. The characteristic of semi-permeable coat can be used in programs of soybean crop breeding to be incorporated in genotypes of high production, aiming at the physiologic quality of the seed. The objectives of this study were: Identify structural differences among the coat of the genotypes CD-202 (permeable) and TP (semi-permeable); obtain fragments of genes distinguishingly expressed among the coat of those genotypes; characterize the physiological quality of soybean seeds produced. Morphologic characterization of the coats was carried out by optic microscope BX 51 with increase of 40x. In order to obtain the fragments of genes distinguishingly expressed among the coats of the two genotypes, the technique cDNA AFLP was used. The physiological quality of the seeds was obtained by germination tests and vigour (accelerated ageing and electrical conductivity). Besides, structural differences were observed among the coats of the two genotypes, however, the genotype TP presented higher thickness in the layers of the epidermis and hypodermis of the coat in relation to the genotype CD 202, beside differences in the format and organization of the cells. Regarding the molecular genetic studies, 47 fragments distinguishingly expressed in the coats in the formation of genotypes CD - 202 and TP, were identified. In agreement with the results, soybean seeds with semi-permeable coat presented better physiological quality in relation to the seeds with permeable coat. / Alguns trabalhos têm evidenciado a existência de genótipos de soja contrastantes para qualidade fisiológica de semente. Tais diferenças podem existir em virtude da presença de sementes com total ou parcial impermeabilidade à penetração de água no tegumento, o que as tornam menos susceptíveis aos danos mecânicos, as adversidades climáticas e a deterioração por umidade. A característica de tegumento semi-permeável pode ser utilizada nos programas de melhoramento de soja para ser incorporada às cultivares de alta produção visando à qualidade fisiológica da semente. Diante do exposto, os objetivos desse estudo foram: identificar diferenças estruturais entre os tegumentos de sementes de soja dos genótipos CD-202 (tegumento permeável) e TP (tegumento semi-permeável); obter fragmentos de genes diferencialmente expressos entre os tegumentos dos dois genótipos; avaliar a qualidade fisiológica das sementes produzidas. A caracterização morfológica dos tegumentos foi realizada através de avaliações em microscópio ótico BX 51 com aumento de 40x. Para obtenção dos fragmentos de genes diferencialmente expressos entre os tegumentos dos dois genótipos, utilizou-se a técnica cDNA AFLP. A avaliação da qualidade fisiológica das sementes foi através dos testes de germinação e vigor (condutividade elétrica e envelhecimento acelerado). Na avaliação morfológica, foram observadas diferenças estruturais entre os tegumentos de semente dos dois genótipos, sendo que, os tegumentos do genótipo TP, apresentaram maior espessura nas camadas da epiderme e hipoderme, além de outras diferenças no formato e organização das células. Com relação aos estudos genéticos moleculares, foram identificados 47 fragmentos de genes diferencialmente expressos entre os tegumentos dos dois genótipos. Na avaliação da qualidade fisiológica das sementes, o genótipo TP apresentou qualidade fisiológica superior em relação ao genótipo CD-202.

Made available in DSpace on 2014-08-20T13:44:44Z (GMT). No. of bitstreams: 1 tese_democrito_amorim_chiesa_freitas.pdf: 404655 bytes, checksum: c873f95da6df2da75ff9fd90ecb3dc2b (MD5) Previous issue date: 2009-03-25 / The need to widen the seeding window of irrigated rice cultivars capable of germinating and producing normal seedlings under low soil temperatures led to the execution of this work, at the laboratory of the Plant Science Department from the Faculty of Agronomy Eliseu Maciel , Federal University of Pelotas and greenhouses at Embrapa s Temperate Climate (CPACT - Pelotas/RS) and Rice & Bean (CNPAF - Goiânia/GO) Experimental Stations. The experimental work focused on the identification of irrigated rice genotypes from an F2 generation tolerant to low soil temperatures at the germination and emergence stages, through the selection of cDNA fragments differentially expressed as the result to low temperature exposure during the germination phase. The procedure consisted of three stages; First stage involved selection, through physiological traits associated to genotype tolerance and sensitivity to germination onset and emergence under low temperature, among the 104 genotypes tested. The Second stage focused on the screening for variability for low temperature tolerance trait among the F2 progeny of a dialellic set of crossings, while the Third stage related to differentially expressed cDNA fragment selection, among embryos from low temperature tolerant and sensitive genotypes. Main results are as follows; a) A large variability for traits expressing tolerance to germination and emergence under low temperatures was determined among the genotypes tested, with BRS Atalanta, LTB 06002, LTB 06012 and LTB 06020 genotypes exhibiting superior germination values, whereas genotypes LTB 06014, Nourim Mochi and Oro exhibited higher emergence values. b) The rice genotypes with high germination values under low temperature conditions did not show the same trend during the emergence stage. c) The cDNA/AFLP technique proved effective in producing and identifying rice gene fragments differentially expressed, under ambient conditions of low temperature during germination. d) Four polymorphic fragments were identified for genotype BRS Firmeza at a temperature of 13°C, thus indicating that four genes were expressed under this condition. / Buscando ampliar a janela de semeadura, com cultivares de arroz irrigado que germinem e desenvolvam plântulas normais em baixa temperatura de solo, realizou-se este trabalho nos laboratórios do Departamento de Fitotecnia da Faculdade de Agronomia Eliseu Maciel da Universidade Federal de Pelotas e nas casas de vegetação da Embrapa Clima Temperado - CPACT (Pelotas/RS) e Embrapa Arroz e Feijão CNPAF (Goiânia/GO). O estudo objetivou identificar genótipos de arroz irrigado tolerantes a baixa temperatura, nos estádios de germinação e emergência, o caráter germinação no frio, na população F2 e selecionar fragmentos de cDNA diferencialmente expressos no frio na germinação. O trabalho foi dividido em três etapas: Primeira etapa, seleção fisiológica, dos genótipos tolerantes e sensíveis ao frio na germinação e na emergência, entre os 104 genótipos estudados, em temperaturas previamente selecionadas; Segunda etapa, observação da existência de variação, na descendência dos cruzamentos dialélicos, para a tolerância ao frio na população F2 , em relação aos pais; Terceira etapa, seleção dos fragmentos de cDNA diferencialmente expressos, entre os embriões de genótipos tolerante e sensível ao estresse abiótico do frio. a) Há variabilidade, no arroz irrigado, quanto à tolerância à baixa temperatura no estádio de germinação e emergência, destacando-se na germinação o BRS Atalanta, LTB 06002, LTB 06012 e LTB 06020, na emergência o LTB 06014, Nourim Mochi e Oro. b) Os genótipos que apresentaram melhor germinação, em baixa temperatura, não mantiveram a superioridade na emergência. c) A técnica de cDNA/AFLP é eficiente para produzir e identificar fragmentos de genes diferencialmente expressos, em genótipos de arroz irrigado durante a germinação, sob estresse de baixas temperaturas. d) No genótipo BRS Firmeza, foram identificados quatro fragmentos polimórficos na temperatura à 13ºC, indicando que quatro genes se expressaram nesta temperatura.

La conductivité hydraulique foliaire (KF) est une composante majeure du transport d’eau dans toute la plante. Dans les feuilles de noyer, la KF est stimulée à la lumière et est étroitement liée à l’accroissement du taux des transcrits d’aquaporines JrPIP2s. Par ailleurs, la corrélation entre la stimulation de la KF et des transcrits d’aquaporines à la lumière, n’est pas générale et dépend de l’espèce. Ici, nous étudions cette corrélation chez cinq espèces forestières (Juglans regia, Fagus sylvitica, Quercus robur, Salix alba et Populus tremula) différant par leur réponse à la lumière. Nous démontrons seulement chez le noyer (Juglans regia), la contribution des deux familles d’aquaporines PIP1s et PIP2s. Afin de mieux comprendre le rôle des JrPIP1s et JrPIP2 dans la réponse à la lumière, nous avons isolé 8 nouvelles isoformes dans les feuilles de noyer et nous avons étudié leurs profils d’expression sur une cinétique lumière. Toutes les isoformes étudiées sont accumulées à la lumière et réprimées à l’obscurité. De plus, la KF est dépendante de la qualité de lumière. Elle est réduite de 65% en absence de lumière bleue. Cette diminution serait liée à l’inhibition des transcrits d’aquaporines. Afin de caractériser les mécanismes moléculaires précoces impliqués dans la modulation de KF par la lumière, l’approche globale cDNA-AFLP a été menée sur des feuilles de noyer sous différentes conditions d’éclairement. Nous obtenons 12000 transcrits différentiels dérivés (TDFs) générés par les 128 couples d’amorces. Parmi les 187 séquences obtenues, 93 d’entre elles ont une fonction putative. Leur classification fonctionnelle montre que les gènes relatifs à la régulation cellulaire représentent environ 58% des TDFs identifiés. Les feuilles exposées à la lumière, montrent des changements dans les voies de : signalisation calcique, protéolyse, trafic vésiculaire et l’expression de divers facteurs de transcription et protéines de régulation. Pour mieux comprendre le rôle potentiel de la signalisation calcique dans la modulation de la KF par la lumière, nous avons étudié l’effet d’un inhibiteur des canaux calciques [LaCl3] et d’un antagoniste de calmoduline [W7] sur la KF et les transcrits des 10 JrPIPs. Comparées aux feuilles témoins, les inhibiteurs calciques provoquent une réduction de la KF et de la majorité des JrPIPs étudiées à la lumière. Nos résultats confirment l’implication du complexe Ca2+ /calmoduline dans la transduction du signal lumineux responsable de la stimulation de la KF et des transcrits d’aquaporines chez le noyer. / Leaf hydraulic conductance (Kleaf) takes a significant part in plant water relations. In walnut leaves, Kleaf was stimulated by light and tightly related to accumulation of JrPIP2s aquaporin transcripts. However, the light effect on Kleaf value is not systematically related to aquaporin regulation. Here we investigated the relationship between light, Kleaf and transcript levels of aquaporin in five species (Juglans regia, Fagus sylvitica, Quercus robur, Salix alba and Populus tremula) differing by the response of their Kleaf to light. Only for walnut leaves, we showed that light-increased Kleaf value is closely related to higher stimulation of both PIP1s and PIP2s aquaporins. To further investigate the involvement of aquporins (JrPIP1s and JrPIP2) in the light Kleaf modulation, 8 new full length aquaporins have been identified in walnut leaves and their expression pattern was monitored. All the aquaporin tested was turned up to be upregulated under light condition and downregulated under darkness. Moreover, we showed that the Kleaf response to light is quality-dependant, since it was reduced of 65% in the absence of blue light. Interstingly, this Kleaf reduction was correlated with a high downregulation of almost all aquaporins tested. To give an insight into the early molecular events involved in the light-induced Kleaf regulation, a large-scale transcriptomic analysis consisting of the cDNA-AFLP procedure was carried out on walnut leaves, kept at different light conditions. We obtained a total of 12,000 transcript-derived fragments (TDFs) by cDNA-AFLP with 128 primer pairs. Reliable sequences were obtained for 187 of these TDFs, and functions were attributed to 93 TDFs through BLAST searches in GenBank databases. Most of the 93 TDFs corresponded to genes encoding proteins involved in cellular regulation (58%). Leaves exposed to light showed changes in the Ca2+-signaling pathway, the ubiquitin-proteasome pathway, vesicle trafficking process and the expression of multiple transcription factors and protein regulators. To progress in understanding of a potential role for calcium signalling in light-modulated Kleaf, Kleaf values and transcript accumulation of 10 JrPIPs were monitored in leaves perfused with either a blocker of calcium channels [LaCl3] or a CaM antagonist [W7]. Compared to control, these Ca2+ -effectors led to a significant reduction in Kleaf and transcripts accumulation of almost all JrPIPs under light conditions. Our results indicate that Ca2+ /calmodulin complex may transduce the light signal required for stimulation of Kleaf and its correlated aquaporin expression.

O carcinoma de célula renal (CCR) subtipo célula clara é o câncer mais letal e prevalente do sistema urinário. A transformação maligna no CCR está possivelmente associada à mudanças no perfil de expressão de oncogenes e genes supressores de tumor, e acredita-se que estas alterações sejam críticas para o desenvolvimento do fenótipo maligno. Para identificar novos genes e vias moleculares associadas à transformação maligna no CCR célula clara, foram analisados perfis de expressão gênica de amostras pareadas de tumor e tecido não tumoral adjacente de 6 pacientes. Foi utilizada uma plataforma de microarrays de cDNA contendo 2.292 sondas mapeando éxons de genes codificadores e 822 sondas de RNAs não-codificadores mapeando em regiões intrônicas. A transcrição intrônica foi detectada em todos os tecidos normais e neoplásicos. Utilizando uma combinação de dois testes estatísticos e uma validação por leave-one-out, foi selecionado um subconjunto de 64 transcritos com expressão significativamente alterada em CCR célula clara em relação ao tecido não tumoral adjacente, estando a maior parte (86%) com expressão diminuída em CCR. Entre os transcritos com expressão diminuída, 49 mapearam em regiões não-traduzidas ou éxons de genes codificadores e 6 mapearam em regiões intrônicas de genes codificadores conhecidos. Os níveis de expressão diminuída de SIN3B, TRIP3, SYNJ2BP e NDE1 (p < 0,02), e de transcritos intrônicos derivados dos loci de SND1 e ACTN4 (p < 0,05), foram confirmados em CCR célula clara por Real-time RT-PCR. Um subconjunto de 25 transcritos se mostrou alterado em 6 amostras adicionais de CCR não célula clara, indicando alterações transcricionais comuns em CCR independentemente do subtipo histológico ou do estado de diferenciação do tumor. Além disso, foi analisado o perfil de metilação dos genes com expressão diminuída em tumor SIN3B, TRIP3, SYNJ2BP e GPX3. Nossos resultados indicam um novo conjunto de candidatos a gene supressor de tumor, que 8 podem desempenhar um papel importante na transformação maligna de células renais normais. / The clear cell subtype of renal cell carcinoma (RCC) is the most lethal and prevalent cancer of the urinary system. The carcinogenesis in RCC is thought to be associated with changes in the expression of several genes, and this alteration in gene expression is believed to be critical to the development of the malignant phenotype. To investigate new genes and molecular pathways associated with malignant transformation in clear cell RCC, gene expression profiles of matched samples of tumor and adjacent non-neoplastic tissue obtained from 6 patients were analysed. A custom-built cDNA microarray platform was used, comprising 2,292 probes that map to exons of genes and 822 probes for noncoding RNAs mapping to intronic regions. Intronic transcription was detected in all normal and neoplastic renal tissues. A subset of 64 transcripts with levels significantly deregulated in clear cell RCC relative to the matched non-tumor tissue, mostly (86%) downregulated in CCR, was

Bean pod mottle virus (BPMV), a member of the genus Comovirus in the family Comoviridae, is widespread in the major soybean-growing areas in the United States. Soybean yield losses of 10-40% have been reported as a consequence of BPMV infection. The complete nucleotide sequences of two strains, K-Ha1 and K-Ho1, were determined. Field isolates of BPMV were classified into two distinct subgroups (I and II) based on slot blot hybridization and sequence analyses. Full-length cDNA clones from which infectious transcripts can be produced were constructed for strains K-G7, K-Ho1 and K-Ha1. Whereas strains K-Ha1 and K-G7 induced mild or moderate symptoms in infected soybean plants, strain K-Ho1 produced very severe symptoms. Symptom severity was mapped to RNA1. Chimeric RNA1 constructs were generated by exchanging full or partial coding regions of the five RNA1-encoded mature proteins between the full-length cDNA clones of the three RNA1s and the resultant transcripts were inoculated onto soybean. The results showed that the coding regions of the protease co-factor (Co-pro) and the putative helicase (Hel) are determinants of symptom severity. Although symptom severity correlated well with accumulation of viral RNA, neither the Co-pro nor Hel protein could be demonstrated as a suppressor of RNA silencing. Furthermore, separate expression of the Co-pro or Hel proteins from a PVX vector induced necrosis on the inoculated leaves of Nicotiana benthamiana. Characterization of BPMV K-Ho1 indicated that it is a diploid reassortant, containing two distinct types of RNA1s and one type of RNA2. Examination of field isolates from various locations in the United States and Canada revealed that diploid reassortants are of frequent occurrence in natural populations of BPMV. The vary severe symptoms induced by BPMV K-Ho1 can be mimicked by inoculation of plants with a mixture of RNA1 transcripts from two distinct strain subgroups and RNA2 transcript from either subgroup. Plants inoculated with a mixture of transcripts containing two types of RNA1 from the same strain subgroup did not produce very severe symptoms. These are due to interactions between two distinct types of RNA1s. At present, no soybean cultivars with resistance to BPMV are commercially available. Therefore, the feasibility of cross protection as an alternative disease management strategy was studied. Two mild strains of BPMV (K-Da1 and K-Ha1), belonging to subgroup II, were tested for their ability to protect infected plants against a severe strain (K-Ho1). Inoculation of the soybean cultivar Essex on the primary leaves with either of the two mild strains conferred complete protection against challenge inoculation with the severe strain K-Ho1, regardless of the timing of challenge inoculation. Cross-protection was evident regardless of whether virions or BPMV-RNA were used as inocula. Cross protection was independent of the soybean cultivar used and method of virus inoculation, sap-inoculation or by the bean leaf beetle, vector of BPMV. Protection was complete and durable.

This dissertation includes three topics. First topic: Regularized estimation in the AFT model with high dimensional covariates. Second topic: A novel application of quantile regression for identification of biomarkers exemplified by equine cartilage microarray data. Third topic: Normalization and analysis of cDNA microarray using linear contrasts.