Thesis (M.A.)--Humboldt State University, 2006. / Includes bibliographical references (leaves 131-133). Also available via Humboldt Digital Scholar.

Thesis (Ph. D.)--Lehigh University, 2006. / Includes vita. Includes bibliographical references (leaves 256-272).

Orientador: Paulo Eduardo Martins Ribolla / Resumo: A deficiência da enzima Glicose-6-Fosfato Desidrogenase (G6PD) é uma anormalidade genética de alta prevalência populacional que resulta em uma menor reatividade do sistema de óxido-redução eritrocitário, geralmente sem repercussões clínicas; estima-se que mais de 300 milhões de pessoas são portadoras dessa alteração. A enzima é expressa em todos os tecidos e catalisa a primeira etapa da Via das Pentoses. Nas hemácias, essa via é de fundamental importância na manutenção do equilíbrio de seu estado redox e a deficiência dessa enzima pode favorecer eventos hemolíticos agudos e crônicos; e em recém nascidos, pode contribuir para o agravamento da icterícia neonatal. O diagnóstico da deficiência baseia-se na atividade enzimática, identificada através de testes quantitativos e qualitativos. Os testes qualitativos limitam-se a agrupar indivíduos em “deficientes” e “não deficientes”, já os métodos quantitativos são mais precisos na inferência dessa atividade. Estas técnicas podem necessitar de repetições dos testes para confirmação de resultados incongruentes. Por outro lado, a variante genética responsável pela deficiência pode ser precisamente reconhecida através de testes de diagnóstico molecular. O presente projeto tem como objetivo desenvolver uma metodologia de identificação molecular da variante G202A, frequentemente encontrada na população brasileira, e realizar uma comparação do custo-benefício com a metodologia de sequenciamento de Sanger. Ao todo, foram analisadas 107 amost... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Glucose-6-phosphate dehydrogenase deficiency is a metabolic enzymatic defect affecting 300 million people worldwide. The enzyme is present in all tissues and catalyses the first reaction in the Pentose Phosphate pathway responsible for maintaining the redox equilibrium in red blood cell. Deficient enzyme may lead to acute and chronic haemolytic anaemia and neonatal jaundice. Diagnosis for G6PD deficiency is based on biochemical quantitative or qualitative tests. Qualitative tests only classifies subjects as “deficient” or “non-deficient”, while quantitative tests are more precise, however both biochemical approches need a confirmative assay to confirm ambiguous results. On the other hand molecular identification for the molecular variants are more accurate and precise. We developed a new molecular assay to identify the G202A molecular variant present at high frequency on Brazilian population and comparer it to Sanger sequencing. One hundred and seven peripheral blood sample were collected on filter paper. DNA extraction were performed followed by G6PD exon 4 amplification and sequencing. On-line tool “Primer1” generated allele-specific primers for TETRA-ARMS genotyping. Twenty two subjects were deficient homozygote, eighty four wild homozygote and one heterozygote. All subjects genotype were confirmed by Sanger sequencing. TETRA-ARMS costs per reaction is three times lower than Sanger sequencing. We conclude that TETRA-ARMS is a suitable protocol to detect G202A mutation on h... (Complete abstract click electronic access below) / Mestre

Typescript. / Bibliography: leaves 184-191. / Photocopy. / Microfilm. / xii, 191 leaves, bound 29 cm

Thesis (M.A.)--Bowling Green State University, 2010. / Document formatted into pages; contains vii, 116 p. Includes bibliographical references.

Made available in DSpace on 2018-08-01T21:35:03Z (GMT). No. of bitstreams: 1 tese_12057_Dissertação_Dalila Avila Silva.pdf: 1086948 bytes, checksum: 30b541bd3051003faada33f9146bf6e5 (MD5) Previous issue date: 2018-02-27 / A Osteogênese Imperfeita (OI) é uma doença que ocorre devido à desordem generalizada do tecido conjuntivo causando principalmente fragilidade óssea. Essa desordem, na maioria das vezes, é causada por mutações nos genes produtores das cadeias do colágeno tipo I, COL1A1 ou COL1A2, embora mutações em novos genes envolvidos na via do metabolismo ósseo têm sido constantemente descobertas. Atualmente, existem 17 genes relacionados com esta doença. Um deles é o gene IFITM5, ainda pouco caracterizado na maioria das populações. Por possuir vários genes causadores de OI, o Next Generation Sequencing (NGS), uma técnica de sequenciamento em larga escala, tem sido amplamente utilizada. Contudo, as mutações identificadas por NGS precisam ser validadas para dar confiabilidade aos resultados. Assim, este trabalhou visou identificar mutações no gene IFITM5 e validar mutações identificadas por NGS nos genes COL1A1 e COL1A2 para caracterizar o padrão de mutações nestes genes em pacientes do Espírito Santo. Este estudo contou com uma amostra inicial de 31 pacientes que foram previamente analisados para outros genes. Desta amostra, 8 indivíduos que apresentaram resultados moleculares inconclusivos foram estudados para o gene IFITM5. Foi detectada a presença da mutação c.-14C>T em um paciente. Esta mutação ocorre na região 5-UTR do gene e é recorrente em várias populações do mundo. A validação de mutações foi realizada em 16 indivíduos que apresentaram alterações genéticas nos genes COL1A1 ou COL1A2 detectadas por NGS. Apenas uma das sequências identificadas por NGS não foi validada. Este estudo confirmou que mutações no gene COL1A1 e COL1A2 são encontradas, em aproximadamente 75% dos pacientes, enquanto que no gene IFITM5 são encontradas mutações em, aproximadamente, 3% dos pacientes com OI do Espírito Santo. Esses resultados poderão auxiliar no desenvolvimento de estratégias de diagnóstico molecular mais eficientes para esta doença.

The main purpose of this study is to analyze the values and results of one year's participation of the Sanger School in a three-year Texas Health Education Program (Texas Extended School and Community Health Program) in order to determine an adequate basis for making recommendations for the further development of this program.

DNA sequence assembly is one of the fundamental areas of bioinformatics. It involves the correct formation of a genome sequence from its DNA fragments ("reads") by aligning and merging the fragments. There are different sequencing technologies -- some support long DNA reads and the others, shorter DNA reads. There are sequence assembly programs specifically designed for these different types of raw sequencing data.<p> This work explores and experiments with these different types of assembly software in order to compare their performance on the type of data for which they were designed, as well as their performance on data for which they were not designed, and on mixed data. Such results are useful for establishing good procedures and tools for sequence assembly in the current genomic environment where read data of different lengths are available. This work also investigates the effect of the presence or absence of quality information on the results produced by sequence assemblers.<p> Five strategies were used in this research for assembling mixed data sets and the testing was done using a collection of real and artificial data sets for six bacterial organisms. The results show that there is a broad range in the ability of some DNA sequence assemblers to handle data from various sequencing technologies, especially data other than the kind they were designed for. For example, the long-read assemblers PHRAP and MIRA produced good results from assembling 454 data. The results also show the importance of having an effective methodology for assembling mixed data sets. It was found that combining contiguous sequences obtained from short-read assemblers with long DNA reads, and then assembling this combination using long-read assemblers was the most appropriate approach for assembling mixed short and long reads. It was found that the results from assembling the mixed data sets were better than the results obtained from separately assembling individual data from the different sequencing technologies. DNA sequence assemblers which do not depend on the availability of quality information were used to test the effect of the presence of quality values when assembling data. The results show that regardless of the availability of quality information, good results were produced in most of the assemblies.<p> In more general terms, this work shows that the approach or methodology used to assemble DNA sequences from mixed data sources makes a lot of difference in the type of results obtained, and that a good choice of methodology can help reduce the amount of effort spent on a DNA sequence assembly project.

DNA sequence assembly is one of the fundamental areas of bioinformatics. It involves the correct formation of a genome sequence from its DNA fragments ("reads") by aligning and merging the fragments. There are different sequencing technologies -- some support long DNA reads and the others, shorter DNA reads. There are sequence assembly programs specifically designed for these different types of raw sequencing data.<p> This work explores and experiments with these different types of assembly software in order to compare their performance on the type of data for which they were designed, as well as their performance on data for which they were not designed, and on mixed data. Such results are useful for establishing good procedures and tools for sequence assembly in the current genomic environment where read data of different lengths are available. This work also investigates the effect of the presence or absence of quality information on the results produced by sequence assemblers.<p> Five strategies were used in this research for assembling mixed data sets and the testing was done using a collection of real and artificial data sets for six bacterial organisms. The results show that there is a broad range in the ability of some DNA sequence assemblers to handle data from various sequencing technologies, especially data other than the kind they were designed for. For example, the long-read assemblers PHRAP and MIRA produced good results from assembling 454 data. The results also show the importance of having an effective methodology for assembling mixed data sets. It was found that combining contiguous sequences obtained from short-read assemblers with long DNA reads, and then assembling this combination using long-read assemblers was the most appropriate approach for assembling mixed short and long reads. It was found that the results from assembling the mixed data sets were better than the results obtained from separately assembling individual data from the different sequencing technologies. DNA sequence assemblers which do not depend on the availability of quality information were used to test the effect of the presence of quality values when assembling data. The results show that regardless of the availability of quality information, good results were produced in most of the assemblies.<p> In more general terms, this work shows that the approach or methodology used to assemble DNA sequences from mixed data sources makes a lot of difference in the type of results obtained, and that a good choice of methodology can help reduce the amount of effort spent on a DNA sequence assembly project.

Background: Lung cancer is the leading cause of cancer-related death and one of the most common cancer types worldwide. Epidermal growth factor receptor (EGFR) has been shown to be an important therapeutic target in non-small cell lung cancer. Kirsten rat sarcoma viral oncogene homologue (KRAS) is a downstream signalling molecule in the EGFR pathway. Lung cancer patients with EGFR mutations respond to tyrosine EGFR inhibitor therapy, in contrast, patients with KRAS mutations do not benefit of such treatment. Methods: This study investigates the frequency of EGFR and KRAS mutations in non-small cell lung cancer patients. Fifty-one lung cancer patients with primary non-small cell lung cancer diagnosed between 1995 and 2005 in the Uppsala-Örebro region were analysed by Sanger sequencing and Pyrosequencing to determine the mutation status of these genes. Results: Five EGFR mutations were found in four patients (8%), two deletions in exon 19, one point mutation in exon 20 and two point mutations in exon 21. KRAS mutations were found in 12 patients (24%), ten codon 12 mutations and two codon 61 mutations. Conclusions: This study confirms previous observations regarding the frequency of EGFR and KRAS mutations in non-small cell lung cancer. Mutations in EGFR and KRAS were mutually exclusive, indicating that both mutations present relevant tumorigenic genomic aberrations.