Submitted by Sara Ribeiro (sara.ribeiro@ucb.br) on 2017-09-08T18:11:30Z No. of bitstreams: 1 OrzenilBonfimdaSilvaJuniorTeseParcial2017.pdf: 781918 bytes, checksum: 8eef627ca550957f6bfa1f46e54c687c (MD5) / Approved for entry into archive by Sara Ribeiro (sara.ribeiro@ucb.br) on 2017-09-08T18:11:38Z (GMT) No. of bitstreams: 1 OrzenilBonfimdaSilvaJuniorTeseParcial2017.pdf: 781918 bytes, checksum: 8eef627ca550957f6bfa1f46e54c687c (MD5) / Made available in DSpace on 2017-09-08T18:11:38Z (GMT). No. of bitstreams: 1 OrzenilBonfimdaSilvaJuniorTeseParcial2017.pdf: 781918 bytes, checksum: 8eef627ca550957f6bfa1f46e54c687c (MD5) Previous issue date: 2017-07-11 / In the last twenty-five years, we have witnessed the wide adoption of DNA markers for the study of genetic variation in many organisms. A DNA marker must have two or more identifiable allelic DNA sequences to be useful. It usually does not have a biological effect, but instead functions as a traceable landmark in the genome, found in a specific location, and transmitted by the standard laws of inheritance from one generation to the next. Its application goes beyond genetic mapping and includes the analysis of genetic diversity, marker-trait association studies, marker assisted selection and, more recently, with the advent of wholegenome sequencing, whole-genome association and genomic selection. Among the several types of DNA sequence polymorphisms that can be used as DNA marker, Single Nucleotide Polymorphisms (SNPs) are the most powerful for large-scale variation analysis. There are vast numbers of SNPs in every genome and they can be typed by methods that have been proven easy to automate. Detection of alternative alleles is rapid and effortless because it is based on well-known polymerase chain reaction and DNA oligomer hybridization assays. Various strategies have been devised to discriminate alleles at a SNP, including fixed DNA arrays technologies, solution hybridization techniques and many sequencing-based genotyping. In our study, we have developed high-throughput DNA marker systems for non-model, highly heterozygous, diploid tree species. We took advantage of the combined power of Next Generation Sequencing (NGS) technologies, well-established highly automated methods of SNP typing and bioinformatics algorithms to perform genome-wide DNA variation analysis. We used whole genome resequencing of pooled individuals to develop a high-density 60K SNP chip for Eucalyptus species (EucHIP60k) providing a 96% genome-wide coverage with 1 SNP/12???20 kbp, and 47,069 SNPs at ??? 10 kb from 30,444 of the 33,917 genes in the Eucalyptus genome. We then used high-density SNP data and whole-genome pooled resequencing to examine the landscape of population recombination (??) and theta (??), assess the extent of linkage disequilibrium (r2) and build the highest density linkage maps for Eucalyptus to date. Chromosome-wide ancestral recombination graphs allowed us to date the split of Eucalytpus grandis (1.7???4.8 million yr. ago) and identify a scenario for the recent demographic history of the species. In a final set of studies, we built the first genome assembly for a Neotropical forest tree, the Pink Ip?? (Handroanthus impetiginosus), a highly-valued keystone timber species. Genome sequence was screened for the development of a targeted-capture sequencing system for SNP genotyping consisting of nearly 24,000 probe sequences. This genotyping system showed flexibility as it allowed the identification of SNPs across different populations of the species in moderate sample sizes. The good genome coverage, consistent Ts/Tv ratio estimated across samples and fair technical reproducibility between replicates, in terms of recall and precision of the SNP calling and accuracy on genotypes, indicate that this genotyping platform can be confidently used to estimate population genetics parameters and carry out population genomics investigations at the genome-wide scale / ***

Submitted by Caroline Xavier (caroline.xavier@pucrs.br) on 2017-05-04T17:15:51Z No. of bitstreams: 1 TES_HENRIQUE_VIEIRA_FIGUEIRO_COMPLETO.pdf: 4680551 bytes, checksum: 8695b78fe6812f4690586975941c4c31 (MD5) / Made available in DSpace on 2017-05-04T17:15:51Z (GMT). No. of bitstreams: 1 TES_HENRIQUE_VIEIRA_FIGUEIRO_COMPLETO.pdf: 4680551 bytes, checksum: 8695b78fe6812f4690586975941c4c31 (MD5) Previous issue date: 2016-03-11 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior - CAPES / Conselho Nacional de Pesquisa e Desenvolvimento Cient?fico e Tecnol?gico - CNPq / In the past 10 years, high throughput sequencing has revolutionized evolutionary biology. With the technical advances that emerged with the genome sequencing of model species, it is now possible to apply these techniques to taxonomic groups without any previously available genetic resources. Complete genome sequencing and reduced representation methods have enabled us to explore deeper evolutionary questions, such as detecting ancient hybridization and signatures of selection on a genomic scale. Among the groups that could benefit from these methods is the Panthera genus. The group is composed by five species (P. onca, P. tigris, P. leo, P. pardus and P. uncia), all of which are large felids that exert important ecological role as apex predators in their habitats. Their low densities, alarming rates of habitat loss and chronic conflict with humans, all of them are threatened with extinction in the wild and thus important targets for conservation. One of the species in this group, the jaguar (P. onca), is the only member of the genus currently present in the Neotropical region, and the focus of our study. The jaguar has a color pattern similar to that of the leopard, but a much more robust constitution, with massive jaws and shorter limbs. The present study aims to characterize for the first time the jaguar genome, and to perform comparative analyses with the genomes from all other Panthera species. In addition, we seek to perform population genomic analyses with Brazilian jaguar populations and search for signatures of divergent selection in different regions. We have sequenced four genomic libraries, with an estimated coverage depth of 84x. The complete genome sequence allowed the annotation of 25,441 genes and the description of other genomic features (e.g. ncRNA, microsatellites, numts). Additionally, we have sequenced the genome of a leopard at low coverage, with an estimated depth of 25x. With the addition of these two genomes, we were able obtain a genomic data set containing all five Panthera species, which was used to perform phylogenetic discordance analyses and to detect signatures of selection using a dataset encompassing 13,143 orthologous genes. We were able to demonstrate the presence of hybridization events during the speciation process of the species, as well as signatures of selection in genes potentially involved in important characteristics of these iconic animals. Among them, the jaguar?s robust build, the social behavior of lions, cold environment adaptations in the snow leopard and the tiger?s stripes. Using an exome capture approach, we performed a population genomics study targeting jaguar populations from different Brazilian biomes. In addition to assessments of genetic diversity and population structure, we detected signals of local adaptation using multiple methods. Among the obtained results is the presence of genes under selection that are related to energetic metabolism in the Amazon, body development in the Pantanal and immunity in the Atlantic Forest. Additionally, we observed several pigmentation-related genes under selection in different biomes. Those genes affect not only pigmentation, but also have pleiotropic effects in development and immunity routes. Overall, these results help to understand the evolutionary processes that have shaped the adaptation of Panthera species, and particularly the jaguar, to the environments where they currently live. / Nos ?ltimos 10 anos, o sequenciamento gen?mico de alto desempenho revolucionou a biologia evolutiva. Com os avan?os gerados pelo sequenciamento do genoma completo de esp?cies modelo, agora ? poss?vel aplicar essas t?cnicas em animais com praticamente nenhum recurso gen?tico dispon?vel. O sequenciamento completo de genomas, bem como o uso de t?cnicas de representa??o reduzida, permitem explorar quest?es evolutivas complexas como, por exemplo, detec??o de hibrida??o e assinaturas de sele??o natural em uma escala gen?mica. Dentre os grupos taxon?micos que podem se beneficiar de tais t?cnicas est? o g?nero Panthera. O grupo ? composto por cinco esp?cies atuais (P. onca, P. tigris, P. leo, P. pardus e P. uncia), todas elas apresentando grande porte e atuando como predadores de topo nos ambientes que ocupam. Devido ?s baixas densidades, alarmante perda de habitat e constantes conflitos com humanos, o n?vel de amea?a em que essas esp?cies se encontram ? preocupante. Dentre as esp?cies do grupo, est? a on?a-pintada (P. onca), ?nica integrante do g?nero na regi?o Neotropical e o principal foco deste trabalho. Nesse sentido, o presente estudo busca caracterizar pela primeira vez o genoma da on?a-pintada, incluindo an?lises comparativas com as outras quatro esp?cies do g?nero. Al?m disso, o trabalho tem como objetivo avaliar as popula??es de on?a no Brasil e buscar assinaturas de sele??o divergente nos biomas que ela ocupa. Para o sequenciamento do genoma da esp?cie, foram utilizadas quatro bibliotecas gen?micas, com uma cobertura estimada de 84x. A sequ?ncia do genoma completo permitiu a anota??o de 25.441 genes e a descri??o de outros componentes do genoma (p.ex. ncRNA, microssat?lites, numts). Adicionalmente, foi sequenciado o genoma de um leopardo (P. pardus) com cobertura estimada de 25x. Com esses dois novos genomas, completou-se um conjunto abrangendo todas as cinco esp?cies do g?nero, permitindo a realiza??o de an?lises de discord?ncia filogen?tica para o grupo e detec??o de sele??o positiva utilizando um conjunto de 13.143 genes ort?logos. Foi poss?vel demonstrar eventos de hibrida??o durante o processo de especia??o das esp?cies do g?nero, bem como sinais de sele??o positiva em genes envolvidos em caracter?sticas que se destacam nos grandes fel?deos. Entre eles, fen?tipos potencialmente afetados por genes sob sele??o incluem o cr?nio e membros robustos da on?a-pintada, o comportamento social no le?o, adapta??o ao frio no leopardo das neves e a presen?a de listras no tigre. Com o uso de captura de exoma, que tem como objetivo o sequenciamento do conjunto de exons da esp?cie, foi poss?vel realizar uma nova avalia??o das caracter?sticas gen?ticas de popula??es de on?a-pintada, bem como a detec??o de assinaturas de adapta??o local. Entre os resultados obtidos est? a presen?a de genes sob sele??o relacionados com metabolismo energ?tico em popula??es da Amaz?nia, adapta??es relacionadas com desenvolvimento corporal no Pantanal e imunidade na Mata Atl?ntica. Adicionalmente, foram observados diversos genes de pigmenta??o com assinaturas de sele??o em diferentes biomas. Esses genes, al?m de afetarem a colora??o dos animais, possuem efeitos pleiotr?picos no desenvolvimento e imunidade da esp?cie. Esses resultados auxiliam no entendimento dos processos evolutivos que moldaram a adapta??o das esp?cies do g?nero, e em especial a on?a pintada, aos ambientes que elas ocupam atualmente.