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SNP discovery, high-density genetic map construction, and identification of genes associated with climate adaptation, and lack of intermuscular bone in tambaqui (Colossoma macropomum) / Descoberta de SNP, construção de mapa genético de alta densidade e identificação de genes associados com adaptação climática e ausência da espinha intermuscular em tambaqui (Colossoma macropomum)Nunes, José de Ribamar da Silva 08 March 2017 (has links)
Tambaqui (Colossoma macropomum) is the largest native Characiform species from the Amazon and Orinoco river basins of South America. Tambaqui farming is growing rapidly in Brazil, its production reached 139.209 tons in 2014, what corresponds to 57.7% of increase compared with 2013. However, few genetic studies of tambaqui are currently available. The tambaqui genetic studies for cultured and wild populations need a holistic approach for a rational action facing ecological and market challenges in aquaculture. Approaches based on genetic studies have provided important tools to understand population dynamics, local adaptation, and gene function to improve selection strategies to be applied in breeding programs. The next-generation sequencing (NGS) allowed a great advance in genomic and transcriptomic approaches, especially related to non-model species. The genotype-by-sequencing (GBS) is one of this approaches based on genome complexity reduction using restriction enzymes (REs). This thesis presents the application of these approaches to provide advances in the genetic background for tambaqui studies. The GBS approach provided a high-density SNPs panel that allowed us to develop the first linkage map, and association studies with environmental variables, local adaptation, and lack of intermuscular bones, both using tambaqui as a model. This work can give us many theoretical references to be applied in genetic breeding programs for tambaqui, allowing a better understanding of genetic processes related to traits of interest in aquaculture. / O tambaqui (Colossoma macropomum) é a maior espécie nativa de Characiforme da América do Sul e é encontrado nas bacias do rio Amazonas e Orinoco. O cultivo do tambaqui está crescendo rapidamente no Brasil, sua produção atingiu 139.209 toneladas em 2014, o que corresponde a 57,7% de aumento em relação a 2013. No entanto, poucos estudos genéticos realizados com o tambaqui estão disponíveis atualmente. Estudos genéticos em tambaqui, tanto em populações cultivadas quanto em populações selvagens, necessitam de uma abordagem holística para uma ação racional frente aos desafios ecológicos e mercadológicos na aquicultura. Abordagens baseadas em estudos genéticos têm fornecido ferramentas importantes para se entender a dinâmica populacional, adaptação local e função gênica visando melhorar as estratégias de seleção a serem aplicadas em programas de melhoramento genético. O sequenciamento de nova geração (NGS) permitiu um grande avanço nas abordagens genômicas e transcriptômicas, especialmente relacionadas a espécies não-modelo. A genotipagem por sequenciamento (GBS) é uma dessas abordagens que utilizam enzimas de restrição (REs) para reduzir a complexidade do genoma. Esta tese apresenta a aplicação desta abordagem objetivando proporcionar avanços significativos nos estudos genéticos de base para tambaqui. A técnica de GBS forneceu um painel de SNPs de alta densidade que nos permitiu desenvolver o primeiro mapa de ligação e estudos de associação com variáveis ambientais, adaptação local e ausência de ossos intermusculares no tambaqui. Este trabalho pode nos dar muitas referências teóricas a serem aplicadas em programas de melhoramento genético do tambaqui, permitindo uma melhor compreensão dos processos genéticos relacionados a traços de interesse na aquicultura.
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Bioinformatics challenges of high-throughput SNP discovery and utilization in non-model organisms2014 October 1900 (has links)
A current trend in biological science is the increased use of computational tools for both the production and analysis of experimental data. This is especially true in the field of genomics, where advancements in DNA sequencing technology have dramatically decreased the time and cost associated with DNA sequencing resulting in increased pressure on the time required to prepare and analyze data generated during these experiments. As a result, the role of computational science in such biological research is increasing.
This thesis seeks to address several major questions with respect to the development and application of single nucleotide polymorphism (SNP) resources in non-model organisms. Traditional SNP discovery using polymerase chain reaction (PCR) amplification and low-throughput DNA sequencing is a time consuming and laborious process, which is often limited by the time required to design intron-spanning PCR primers. While next-generation DNA sequencing (NGS) has largely supplanted low-throughput sequencing for SNP discovery applications, the PCR based SNP discovery method remains in use for cost effective, targeted SNP discovery. This thesis seeks to develop an automated method for intron-spanning PCR design which would remove a significant bottleneck in this process. This work develops algorithms for combining SNP data from multiple individuals, independent of the DNA sequencing platforms, for the purpose of developing SNP genotyping arrays. Additionally, tools for the filtering and selection of SNPs will be developed, providing start to finish support for the development of SNP genotyping arrays in complex polyploids using NGS.
The result of this work includes two automated pipelines for the design of intron-spanning PCR primers, one which designs a single primer pair per target and another that designs multiple primer pairs per target. These automated pipelines are shown to reduce the time required to design primers from one hour per primer pair using the semi-automated method to 10 minutes per 100 primer pairs while maintaining a very high efficacy. Efficacy is tested by comparing the number of successful PCR amplifications of the semi- automated method with that of the automated pipelines. Using the Chi-squared test, the semi-automated and automated approaches are determined not to differ in efficacy.
Three algorithms for combining SNP output from NGS data from multiple individuals are developed and evaluated for their time and space complexities. These algorithms were found to be computationally efficient, requiring time and space linear to the size of the input. These algorithms are then implemented in the Perl language and their time and memory performance profiled using experimental data. Profiling results are evaluated by applying linear models, which allow for predictions of resource requirements for various input sizes. Additional tools for the filtering of SNPs and selection of SNPs for a SNP array are developed and applied to the creation of two SNP arrays in the polyploid crop Brassica napus. These arrays, when compared to arrays in similar species, show higher numbers of polymorphic markers and better 3-cluster genotype separation, a viable method for determining the efficacy of design in complex genomes.
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SNP discovery, high-density genetic map construction, and identification of genes associated with climate adaptation, and lack of intermuscular bone in tambaqui (Colossoma macropomum) / Descoberta de SNP, construção de mapa genético de alta densidade e identificação de genes associados com adaptação climática e ausência da espinha intermuscular em tambaqui (Colossoma macropomum)José de Ribamar da Silva Nunes 08 March 2017 (has links)
Tambaqui (Colossoma macropomum) is the largest native Characiform species from the Amazon and Orinoco river basins of South America. Tambaqui farming is growing rapidly in Brazil, its production reached 139.209 tons in 2014, what corresponds to 57.7% of increase compared with 2013. However, few genetic studies of tambaqui are currently available. The tambaqui genetic studies for cultured and wild populations need a holistic approach for a rational action facing ecological and market challenges in aquaculture. Approaches based on genetic studies have provided important tools to understand population dynamics, local adaptation, and gene function to improve selection strategies to be applied in breeding programs. The next-generation sequencing (NGS) allowed a great advance in genomic and transcriptomic approaches, especially related to non-model species. The genotype-by-sequencing (GBS) is one of this approaches based on genome complexity reduction using restriction enzymes (REs). This thesis presents the application of these approaches to provide advances in the genetic background for tambaqui studies. The GBS approach provided a high-density SNPs panel that allowed us to develop the first linkage map, and association studies with environmental variables, local adaptation, and lack of intermuscular bones, both using tambaqui as a model. This work can give us many theoretical references to be applied in genetic breeding programs for tambaqui, allowing a better understanding of genetic processes related to traits of interest in aquaculture. / O tambaqui (Colossoma macropomum) é a maior espécie nativa de Characiforme da América do Sul e é encontrado nas bacias do rio Amazonas e Orinoco. O cultivo do tambaqui está crescendo rapidamente no Brasil, sua produção atingiu 139.209 toneladas em 2014, o que corresponde a 57,7% de aumento em relação a 2013. No entanto, poucos estudos genéticos realizados com o tambaqui estão disponíveis atualmente. Estudos genéticos em tambaqui, tanto em populações cultivadas quanto em populações selvagens, necessitam de uma abordagem holística para uma ação racional frente aos desafios ecológicos e mercadológicos na aquicultura. Abordagens baseadas em estudos genéticos têm fornecido ferramentas importantes para se entender a dinâmica populacional, adaptação local e função gênica visando melhorar as estratégias de seleção a serem aplicadas em programas de melhoramento genético. O sequenciamento de nova geração (NGS) permitiu um grande avanço nas abordagens genômicas e transcriptômicas, especialmente relacionadas a espécies não-modelo. A genotipagem por sequenciamento (GBS) é uma dessas abordagens que utilizam enzimas de restrição (REs) para reduzir a complexidade do genoma. Esta tese apresenta a aplicação desta abordagem objetivando proporcionar avanços significativos nos estudos genéticos de base para tambaqui. A técnica de GBS forneceu um painel de SNPs de alta densidade que nos permitiu desenvolver o primeiro mapa de ligação e estudos de associação com variáveis ambientais, adaptação local e ausência de ossos intermusculares no tambaqui. Este trabalho pode nos dar muitas referências teóricas a serem aplicadas em programas de melhoramento genético do tambaqui, permitindo uma melhor compreensão dos processos genéticos relacionados a traços de interesse na aquicultura.
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