Developing omics and cell culture resources in non-model organism for their application in elucidating complex trait evolution in natural populations.
Type of DegreePhD Dissertation
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In the past, researchers chose models organisms to answer research questions based on their simplicity in morphology, domestication, and/or life history traits. Now, with high throughput sequencing (HTS) rapidly becoming cheaper, more investigations are tractable even with relatively small budgets. This is important because there are many complex and system-specific questions about natural populations and natural phenomenon that cannot be answered with traditional model organisms. Further, we cannot refine our understanding of life and biodiversity in a small unrepresentative subset of model organisms. The goal of this work is the generation of omics and cell culture resources for research on Daphnia and Anolis genera, respectively. I discuss their applicability for investigations of natural populations in the context of conservation and organisms themselves. Chapters One & Two develop omics resources in Daphnia systems, which are highly tractable evolutionary and ecotoxicology models. Chapter One, a published manuscript in G3, describes the generation of reference-guided draft genome assemblies for two strains of D. pulicaria with differential responses to toxic algal blooms. These assemblies with a high-quality curation of genes can be used for several downstream investigations, including the exploration of differential gene expression in response to algal toxins and identification of sequence variants associated with toxin resistance. Chapter Two diverts focus to the technical side of omics investigations where I explore 18 different combinations of RNA-seq tools for DGE analysis using a computationally tractable caloric restriction data set from D. pulex. I discuss the variation in biological or functional results due to the different tool combinations and explore variation at each step of the pipelines. The work presented in this chapter is the basis for three manuscripts in development: (1) the current chapter to be expanded and contrasted across species; (2) the biological inferences from the RNA-seq results in conjunction with phenotypic data; and (3) the code used for these comparisons as a detailed tutorial for instructors to teach these analyses using a non-model system. My final chapter details the generation of cellular resources for testing in silico or omics generated functional predictions. I develop primary and early passage cell lines for > 100 lizards from the Anolis genus, providing methods for establishment and validation of reptile cell lines. I discuss their applicability in the context of studying protected and/or cryptic species in a dish!