DE LA TESIS:Fermat, motivado por la lectura de Diofanto, conjeturó que todo entero primo congruente con 1 módulo 4 es representable de manera única, salvo el signo y el orden, como suma de dos cuadrados; también propuso que todo entero positivo es suma de cuatro cuadrados. Estos resultados fueron probados por Euler y Lagrange. La caracterización de los enteros representables como suma de tres cuadrados fue dada por Legendre. Gauss dio el número total de representaciones primitivas de un entero "n" como suma de tres cuadrados, en función del número de clases de formas cuadráticas binarias de discriminante -"n". El cálculo del número de representaciones de un entero como suma de 2, 4, 6 y 8 cuadrados fue llevado a cabo por Jacobi, mediante la teoría de funciones elípticas. Las fórmulas correspondientes se obtienen igualando coeficientes en ciertas identidades satisfechas por su función "theta". Las investigaciones de Jacobi fueron proseguidas por Liouville y Ramanujan, entre otros, y en ellas se encuentra uno de los orígenes del estudio de las llamadas formas modulares de peso entero. En general, el cálculo del número de representaciones de un entero por una forma cuadrática, entera, concreta es muy complicado y resulta imposible obtener formulas exactas que expresen dicho número. Para paliar este inconveniente, Gauss, en el caso de formas cuadráticas binarias, introdujo el concepto de género; este fue convenientemente extendido por Eisenstein, Smith y Minkowski a formas de un número mayor de variables. Siegel en 1935 dio formulas para una media, convenientemente ponderada, del número de representaciones de un entero por todas las formas que integran un género. El estudio por vía analítica, análogo al de Jacobi, del número de representaciones de un entero como suma de un número impar de cuadrados fue comenzado por Hardy y Mordell, y conduce al estudio de las formas modulares de peso semientero. Este último concepto puede decirse que no ha sido completamente clarificado hasta los trabajos de Shimura de 1973. El caso de tres variables es el más delicado; hasta 1984, en el trabajo de Schulze-Pillot, no se ha probado que la serie "theta" del genero de una forma cuadrática ternaria es una serie de Eisenstein. En esta memoria nos ocupamos del siguiente problema relativo a las sumas de tres cuadrados:Dado un entero "n", hallar el valor de "e" máximo del cual se puede afirmar que existe una representación de "n" como suma de tres cuadrados = x(2 / 1) + x(2 / 2) + x (2 / 3), con "e" sumandos primos con "n"Este problema, aparte de su interés intrínseco, ha sido motivado por su conexión con la búsqueda de enteros "n" para los cuales toda extensión central del grupo alternado A(n) es grupo de Galois sobre "Q". La única referencia existente en la literatura de un problema análogo al que nos ocupa es el resultado elemental de Catalan, de 1880, de que toda potencia de 3 es suma de tres cuadrados primos con 3. La presente memoria está dividida en seis capítulos: En el primer capítulo se hace la presentación del problema, diciéndose de él todo lo que se puede mediante métodos elementales. Se detalla asimismo su relación con el problema inverso de la teoría de Galois, antes mencionado. En el segundo capítulo se dan fórmulas que, de todas las representaciones de un entero como suma de tres cuadrados, descuentan aquellas que tienen "k" términos no primos con "n", para "k" = 1, 2, 3. Como estas formulas son de evaluación imposible, se definen a su vez, en este capítulo, unas fórmulas en "media" que aproximan a las primeras y son evaluables. El estudio de estas últimas se lleva a cabo en los capítulos III y IV. En el capítulo V se estudia el error cometido en la utilización de las formulas en "media" en vez de las fórmulas exactas. Todo ello permite en el capítulo VI dar una respuesta al problema.

No description available.

Microsatellites are strings of short DNA motifs (≤6 bp) repeated in tandem across genomes of both prokaryotes and eukaryotes. In 20 years, they became popular genetic markers, successfully employed in the field of genetic mapping and gene hunting, as well as to address various biological questions at the individual, family, population and species level. However, evolutionary and demographic inferences from microsatellite polymorphism are hampered by controversy and ambiguity in the mutational processes of microsatellite sequences. Drawing on new data from genome projects, I review in Chapter 1 the concept of a microsatellite life cycle, which hypothesizes that microsatellites follow a life cycle from birth, through expansion, contraction, death and potentially resurrection. To document and understand this integrative concept of evolution, which could help improve current models of microsatellite evolution, there is an implicit need to study the evolution of microsatellites above the species level. A prerequisite of such comparative studies is therefore to find microsatellite loci that are conserved between different species. The near or full completion of many vertebrate genomes and their alignment against one another offer the ultimate approach to find genomic elements conserved over a large evolutionary scale. In Chapter 2, I present a new comprehensive method to find conserved microsatellites in whole genomes. Using the multiple-alignment of the human genome against those of 11 mammalian and five non-mammalian vertebrates, I examine the genomewide conservation of microsatellites, and challenge the general assumption that microsatellites are too labile to be maintained in distant species. In Chapter 3, I present similar results using the alignment of the newly sequenced platypus genome against those of three mammals, the chicken and the lizard, and incorporate these data into the framework created by the 17-genome analysis. This enlarged dataset was ground for attempting to reconstruct a vertebrate phylogeny from the presence/absence of microsatellites in the different genomes. Maximum parsimony analyses resulted in a tree much similar to that of the current view of the vertebrate phylogeny, while Bayesian analyses showed some discrepancies. This work opens a way for novel theoretical developments regarding the inference of ancestral states of microsatellites. In Chapter 4, I show how knowledge on conserved microsatellite sites can help for the development of a set of comparative primers useful across the Mammalia; implementing a similar protocol, nine conserved dinucleotide repeats were genotyped in 20 unrelated individuals of 18 species (nine sister species) encompassing the mammalian phylogeny, including marsupials and monotremes, and four microsatellites were sequenced in 4 individuals per species. My results emphasize conserved microsatellites as a new resource for genetic mapping and population studies. Finally, in Chapter 5, I recount the unexpected extent of structural change among mammalian orthologous microsatellites, including change of complexity, motif replacement and overall length variability. Altogether, these findings provide a comprehensive framework that may help in many areas of research, including molecular ecology, genome mapping, population genetics, and genome and microsatellite evolution.

Thesis (Ph. D.)--University of Alabama at Birmingham, 2006. / Title from first page of PDF file (viewed Feb 15, 2008). Includes bibliographical references.

Thesis (Ph.D.)--University of Alabama at Birmingham, 2007. / Title from PDF title page (viewed on Sept. 16, 2009). Includes bibliographical references.

Made available in DSpace on 2012-05-07T14:40:40Z (GMT). No. of bitstreams: 2 license.txt: 1748 bytes, checksum: 8a4605be74aa9ea9d79846c1fba20a33 (MD5) 000021.pdf: 7148485 bytes, checksum: d3a3aca355db988afba0a31f293e9c1e (MD5) Previous issue date: 2008 / O diagnóstico precoce da leishmaniose visceral (LV) é importante para evitar danos severos que podem levar o paciente à morte. Neste contexto, os métodos moleculares vêm sendo desenvolvidos com destaque para a tecnologia da reação em cadeia da polimerase (PCR). Recentemente, a PCR apresentou um significativo avanço em sua tecnologia; é a PCR quantitativa em tempo real (qPCR). Objetivouse desenvolver e avaliar um sistema baseado em qPCR para o diagnóstico da infecção por Leishmania infantum em cães, bem como efetuar uma análise comparativa com a PCR convencional utilizada no Serviço de Referência em Leishmanioses de Pernambuco. Com base na seqüência NCBI Z35273.1 de L. infantum disponível no BLAST-NCBI foram desenhados primers específicos para o complexo L. donovani. A combinação dos primers gerou sistemas de detecção, sendo o sistema Linf 1 B o mais promissor. A curva-padrão foi gerada resultando em limite de detecção de 10 fg de DNA genômico de L. infantum (7x10-2 parasitas), e = 0.9417, R2= 0.931 e slope= -3.47. O sistema desenvolvido foi avaliado em sangue de cães positivos e negativos para leishmaniose visceral canina, apresentando sensibilidade de 100 por cento e especificidade de 83,33 por cento. A análise comparativa com o PCR convencional mostrou que, utilizando-se amostras de sangue, o qPCR é mais sensível (sensibilidade PCR= 23,8 por cento, qPCR= 100 por cento). Em relação à especificidade, apesar da PCR convencional ter apresentado valor de 100 por cento, as análises por meio dos intervalos de confiança e o teste 2? mostraram que os dois testes (PCR convencional e qPCR) são equivalentes. Desta forma, conclui-se que os resultados obtidos para o sistema de qPCR, em amostra de sangue de cães e, a análise comparativa com a PCR convencional (RV1/RV2), sugerem sua utilização nas rotinas de diagnóstico da infecção por L. infantum

La métagénomique ciblée, étude de la composition et de la diversité des communautés microbiennes présentes dans différents échantillon biologiques sur la base d'un marqueur génomique, a connu un véritable essor lors de cette dernière décennie grâce à l'arrivée du séquençage haut-débit. Faisant appel à des outils de biologie moléculaire et de bioinformatique, elle a été à l’origine de substantiels progrès dans les domaines de l’évolution et de la diversité microbienne. Cependant, de nouvelles problématiques sont apparues avec le séquençage haut-débit : la génération exponentielle de données soulève des problèmes d'analyse bioinformatique, qui doit être adaptée aux plans d'expérience et aux questions biologiques associées. Cette thèse propose des solutions d'amélioration des études de métagénomique ciblée par le développement d'outils et de méthodes innovantes, apportant une meilleure compréhension des biais d'analyse inhérents à de telles études, et une meilleure conception des plans d'expérience. Tout d'abord, une expertise du pipeline d'analyse utilisé en production sur la plate-forme PEGASE-biosciences a été menée. Cette évaluation a révélé la nécessité de mettre en place une méthode d'évaluation formelle de pipelines d'analyses de données de métagénomique ciblée, qui a été développée sur la base de données simulées et réelles, et de métriques d'évaluation adaptées. Cette méthode a été utilisée sur plusieurs pipelines d'analyse couramment utilisés par la communauté, tout comme sur de nouvelles approches d'analyse jamais utilisées dans un tel contexte. Cette évaluation a permis de mieux comprendre les biais du plan d'expérience qui peuvent affecter les résultats et les conclusions biologiques associées. Un de ces biais majeurs est le choix des amorces d'amplification de la cible ; un logiciel de design d'amorces adaptées au plan d'expérience a été spécifiquement développé pour minimiser ce biais. Enfin, des recommandations de montage de plan d'expérience et d'analyse ont été émises afin d'améliorer la robustesse des études de métagénomique ciblée. / Targeted metagenomics is the study of the composition of microbial communities in diverse biological samples, based on the sequencing of a genomic locus. This application has boomed over the last decade thanks to the democratisation of high-throughput sequencing, and has allowed substantial progress in the study of microbial evolution and diversity. However, new problems have emerged with high-throughput sequencing : the exponential generation of data must be properly analyzed with bioinformatics tools fitted to the experimental designs and associated biological questions. This dissertation provides solutions to improve targeted metagenomics studies, by the development of new tools and methods allowing a better understanding of analytical biases, and a better design of experiments. Firstly, an expert assessment of the analytical pipeline used on the PEGASE-biosciences plateform has been performed. This assessment revealed the need of a formal evaluation method of analytical pipelines used for targeted metagenomics analyses. This method has been developed with simulated and real datasets, and adequate evaluation metrics. It has been used on several analytical pipelines commonly used by the scientific community, as well as on new analytical methods which have never been used in such a context before. This evaluation allowed to better understand experimental design biases, which can affect the results and biological conclusions. One of those major biases is the design of amplification primers to target the genomic locus of interest. A primer design software, adaptable to different experimental designs, has been specifically developed to minimize this bias. Finally, analytical guidelines and experimental design recommendations have been formulated to improve targeted metagenomics studies.

Two species of Cryptosporidium are routinely found in pigs: Cryptosporidium suis and Cryptosporidium pig genotype II. Identification of Cryptosporidium species and genotypes currently relies on molecular methods such as polymerase chain reaction (PCR) followed by restriction fragment lenght polymorphism (RFLP) or gene sequencing. However their applications are limited in identification of mixed infections. To overcome this problem, novel species specific primers were developed in this study. A total of 457 pig fecal samples were collected and examined using microscopy and molecular tools including PCR-RFLP, species or genus specific nested PCR and sequencing. Of these, 12.8 % were microscopicaly positive for oocysts presence and 36.5 % using molecular methods. While PCR-RFLP with genus specific primers revealed 1 case of C. suis and Cryptosporidium pig genotype II mixed infection only, nested PCR with species specific primers identified 41 cases of mixed infections. Our results showed that C. suis is infectious for all age categories of pigs and Cryptosporidium pig genotype II has been found in animals older than 6 weeks of age. Morphometric analysis proved oocyst size difference between both pig specific Cryptosporidium spp. Histological examination revealed that Cryptosporidium pig genotype II infects epithelia of both small and large intestine.

Made available in DSpace on 2014-08-20T13:32:55Z (GMT). No. of bitstreams: 1 dissertacao_fabiana_fonseca_santos.PDF: 358501 bytes, checksum: 566ce94854cb74cc7a2d26d876462a45 (MD5) Previous issue date: 2010-08-17 / The understanding of the genetic structure of the oat species can be obtained with the use of microsatellite molecular markers which represent powerful tools in the investigation of genetic parameters. This study aimed to search the identification and characterization of microsatellite markers occurrence in oat through bioinformatics, describing the possible role of individual genes based on the genome of rice and Arabidopsis thaliana. For the development and validation of EST (Expressed Sequence Tags) primers, sequences were obtained from a public database. A total of 7632 oat contigs were obtained after eliminating the redundancy. To characterize SSR (simple sequence repeats) loci, the SSRLocator program was used. For the design of primers, the module containing the program Primer3 was used. A total of 58 primers were found, from which ten were selected for validation by polymerase chain reaction. Ten genotypes were used to assess polymorphism detection ability of primers. Polymorphism was observed in 80% of primers, whereas the rest showed monomorphic bands. The high genetic diversity revealed by microsatellite markers in this study may reflect a high genetic diversity within the germplasm of Avena sativa. / O conhecimento da estrutura genética de diversas espécies de aveia pode ser obtido com o uso de marcadores moleculares microssatélites, os quais representam poderosa ferramenta na averiguação de parâmetros genéticos. Este trabalho teve como objetivo buscar a identificação e a caracterização da ocorrência de marcadores microssatélites em aveia através da bioinformática, descrevendo a possível função de cada um dos genes, com base no genoma do arroz e da Arabidopsis thaliana. Visando o desenvolvimento e a validação de primers, foram obtidos a partir de um banco de dados público de EST (Expressed Sequence Tag) de aveia um total de 7.632 contigs que tiveram eliminada sua redundância, e para caracterização de locos SSR (Simple Sequence Repeat) foi utilizado o programa SSRLocator. Para o desenho de iniciadores foi utilizado no SSRLocator o módulo contendo o programa Primer3 sendo encontrados 58 primers, dos quais dez foram selecionados para a validação através da reação em cadeia da polimerase. Dez genótipos de aveia foram avaliados quanto ao nível de polimorfismo. Foi verificado polimorfismo em 80% dos primers, sendo que, o restante apresentou bandas monomórficas. A alta diversidade gênica revelada pelos marcadores microssatélites neste estudo possivelmente reflete uma alta diversidade genética dentro do germoplasma de Avena sativa.

In an increasingly energy-hungry world, our capacity to meet the heightened energy demands of the future has become a pressing matter. The most urgent of these concerns are tied to the accessibility of petroleum. Various experts have proselytized both the imminent arrival of peak oil production rates and the ensuing decline of those rates thereafter. And to that end, the development of novel and advanced oil exploration methodologies has become almost as important as finding the sources of oil themselves. The soils above petroleum reservoirs play host to various communities of alkane- oxidizing bacteria that can utilize the natural gas emitted by the reservoirs as a source of carbon and energy. While methane can originate from non-petroleum sources, the only natural sources of propane and butane are oil and gas fields. The increased presence of propane and butane-oxidizing bacteria in a given soil sample is used by oil prospectors as an accurate indicator of a proximal petroleum reservoirs. For over a century, cell counts and hydrocarbon metabolic rates have been the metrics used to determine the presence of hydrocarbon-oxidizing microbes. These methods require weeks to complete. Here, we have developed a set of DNA primers for a much more rapid detection of hydrocarbon-oxidizing microbes through PCR amplification - for the chief purpose of petroleum exploration. Each primer’s design is based on a nucleotide sequence alignment of seven prmA and bmoX genes from seven organisms, which encode the large hydroxylase subunit of propane monooxygenase and the alpha hydroxylase subunit of butane monooxygenase respectively. These monooxygenases are the enzymes responsible for the initiation of propane and butane catabolism. Optimization of PCR with this primer set was accomplished using DNA extracted from known butane and propane oxidizers as positive controls, and methane and toluene oxidizers as negative controls. PCR products recovered from cultures of butane-oxidizing and propane-oxidizing bacteria, and soil samples, were sequenced. Phylogenetic trees were constructed from the sequencing data to confirm the accuracy of amplification. We demonstrate the use of PCR and agarose gel electrophoresis to detect hydrocarbon-oxidizing bacteria in culture and in complex microbial soil communities. Detection limits were elucidated through two different experiments. Potential avenues of advancements include narrowing specificity by selectively removing primer degeneracies, the use of additional positive and negative controls and the adaptation of the primers to a qPCR TaqMan assay.