Thesis (Ph.D.)--University of California, San Francisco, 2009. / Source: Dissertation Abstracts International, Volume: 70-04, Section: B, page: 2039. Adviser: Didier Y.R. Stainier.

<p> When challenged by difficult biological samples, the forensic analyst is far more likely to obtain useful data by sequencing the human mitochondrial DNA (mtDNA). Nextgeneration sequencing (NGS) technologies are currently being evaluated by the Forensic Science Program at Western Carolina University for their ability to reliably detect lowlevel variants in mixtures of mtDNA. The sequence profiles for twenty individuals were obtained by sequencing amplified DNA derived from the mitochondrial hypervariable (HV) regions using Sanger methods. Two-person mixtures were then constructed by mixing quantified templates, simulating heteroplasmy at discrete sites and in defined ratios. Libraries of unmixed samples, artificial mixtures, and instrument controls were prepared using Illumina^&reg; Nextera^&reg; XT and deep-sequenced on the Illumina^&reg;MiSeq&trade;. Analysis of NGS data using a novel bioinformatics pipeline indicated that minor variants could be detected at the 5, 2, 1, and 0.5% levels of detection. Additional experiments which examined the occurrence of sequence variation in hair tissue demonstrates that a considerable amount of sequence variation can exist between hairs and other tissues derived from a single donor. </p>

<p> The Archaea represent a fascinating domain of life where each species comprises a hybrid of bacterial and eukaryal features. Unfortunately, there have been few investigations of the Archaea and many fundamental questions remain regarding their biochemistry, genetics, and genomics. One reason for this is that few archaea are amenable to detailed experimental analysis; however, a few halophilic archaea (haloarchaea) can be easily manipulated in the laboratory and contain a sequenced genome, allowing for bioinformatics studies, such as recording changes in the transcriptomes. Haloarchaea are found wherever seawater is concentrated above 2 M NaCl and contain a similarly high concentration of salts internally, without producing compatible solutes. They exhibit a variety of novel molecular characteristics, including acidic proteins that resist the denaturing effects of salts, and DNA repair systems that minimize the deleterious effects of desiccation and intense solar radiation. In addition, haloarchaea are metabolically versatile and respond to a wide variety of environmental signals, including extremes of radiation, salinity, temperature, heavy metals, pollutants, and pH by modulating the activity of key genes. </p><p> Of the above naturally occurring stresses, all have been previously studied but one, pH. Haloarchaea are routinely isolated from both acidic and alkaline saline lakes; however, their ability to thrive under these conditions still remains an unexplored mystery. Therefore, I have endeavored to unlock a few of these secrets by growing sequenced haloarchaeal strains at extremes of pH and cataloging the changes in their transcriptomes compared to growth at optimal pH. For my experiments, RNA was isolated at the end of logarithmic growth and labeled cDNA was generated and hybridized to custom-designed microarray slides that I designed. Changes in the transcriptomes were cataloged and compared to each other as well as to changes from previously studied bacteria. The results from my experiments showed that haloarchaea are primarily responding to extremes of pH in a manner similar to known bacterial cells, making these responses similar across two domains of life. However, there are a few differences, compared to bacteria, due to the unique nature of the Archaea.</p>

<p> In the practice of resource management conservation, it is common to introduce new members into small inbred populations in order to increase genetic diversity and reduce the negative effects of inbreeding. Although fitness often increases in the F1 generation immediately following intrapopulation hybridization, few studies have focused on the long-term fitness of such populations. The model organisms <i>Drosophila melanogaster</i> and <i> D. simulans</i> were used to investigate if heterosis, an increase in the fitness in the hybrid population, or outbreeding depression, a decline in the fitness in the hybrid population, occurs over an eight generation period. At each generation, a series of fitness related assays were performed to assess relative fitness. Results of this study were mixed. At best, only some hybridized populations showed slightly higher, but often nonsignificant increases in fitness, never in more than one assay. The lack of consistent strong persistent heterosis suggests that conservation efforts such as habitat preservation maybe be a better use of effort and money for assisting endangered species. </p>

<p> Evolutionary forces such as recombination, demography, and selection can shape patterns of genetic diversity within populations and contribute to phenotypic variation. While theoretical models exist for each of these forces independently, mathematically modeling their joint impact on patterns of genetic diversity remains very challenging. Fortunately, it is possible to perform forward-in-time computer simulations of DNA sequences that incorporate all of these forces simultaneously. Here, I show that there are trade-offs between computational efficiency and accuracy for simulations of a widely investigated model of recurrent positive selection. I develop a theoretical model to explain this trade-off, and a simple algorithm that obtains the best possible computational performance for a given error tolerance. I then pivot to develop a framework for simulations of human DNA sequences and genetically complex phenotypes, incorporating recently inferred demographic models of human continental groups and selection on genes and non-coding elements. I use these simulations to investigate the power of rare variant association tests in the context of rampant selection and non-equilibrium demography. I show that the power of rare variant association tests is in some cases quite sensitive to underlying assumptions about the relationship between selection and effect sizes. This work highlights both the challenge and the promise of applying forward simulations in genetic studies that seek to infer the parameters of evolutionary models and detect statistical associations.</p>

Thesis (Ph.D.)--University of California, San Francisco, 2005. / Source: Dissertation Abstracts International, Volume: 66-12, Section: B, page: 6369. Adviser: Nigel Killeen.

Thesis (Ph.D.)--University of California, San Francisco, 2007. / Source: Dissertation Abstracts International, Volume: 68-07, Section: B, page: 4219. Adviser: Didier Y.R. Stainier.

Thesis (Ph.D.)--University of California, San Francisco, 2008. / Source: Dissertation Abstracts International, Volume: 68-12, Section: B, page: 7754. Adviser: Peter K. Walter.

Thesis (Ph.D.)--University of California, San Francisco, 2007. / Source: Dissertation Abstracts International, Volume: 68-04, Section: B, page: 2051. Adviser: Dean Sheppard.

Thesis (Ph.D.)--University of California, San Francisco, 2005. / Source: Dissertation Abstracts International, Volume: 66-12, Section: B, page: 6399. Adviser: Didier Y. R. Stainier.