The soil-nematode Caenorhabditis elegans survives oxygen deprivation (anoxia < 0.001 kPa of O2, 0% O2) by entering into a state of suspended animation during which cell cycle progression at interphase, prophase and metaphase stage of mitosis is arrested. I conducted cell biological characterization of embryos exposed to various anoxia exposure times, to demonstrate the requirement and functional role of spindle checkpoint gene san-1 during brief anoxia exposure. I conducted a synthetic lethal screen, which has identified genetic interactions between san-1, other spindle checkpoint genes, and the kinetochore gene hcp-1. Furthermore, I investigated the genetic and cellular mechanisms involved in anoxia-induced prophase arrest, a hallmark of which includes chromosomes docked at the nuclear membrane. First, I conducted in vivo analysis of embryos carried inside the uterus of an adult and exposed to anoxic conditions. These studies demonstrated that anoxia exposure prevents nuclear envelope breakdown (NEBD) in prophase blastomeres. Second, I exposed C. elegans embryos to other conditions of mitotic stress such as microtubule depolymerizing agent nocodazole and mitochondrial inhibitor sodium azide. Results demonstrate that NEBD and chromosome docking are independent of microtubule function. Additionally, unlike anoxia, exposure to sodium azide causes chromosome docking in prophase blastomeres but severely affects embryonic viability. …

The primary objectives of this project were: 1. to develop improved methods for extraction of DNA from human skeletal remains, 2. to improve STR profiling success of low-copy DNA samples by employing whole genome amplification to amplify the total pool of DNA prior to STR analysis, and 3. to improve STR profiling success of damaged DNA templates by using DNA repair enzymes to reduce the number/severity of lesions that interfere with STR profiling. The data from this study support the following conclusions. Inhibitory compounds must be removed prior to enzymatic amplification; either during bone section pretreatment or by the DNA extraction method. Overall, bleach outperformed UV as a pretreatment and DNA extraction using silica outperformed microconcentration and organic extraction. DNA repair with PreCR™ A outperformed both whole genome amplification and repair with PreCR™ T6. Superior DNA extraction results were achieved using the A6 PMB columns (20 ml capacity column with 6 layers of type A glass fiber filter), and DNA repair with PreCR™ A led to an overall improvement in profile quality in most cases, although whole genome amplification was unsuccessful. Rapid, robust DNA isolation, successful amplification of loci from the sample-derived DNA pool, and an elimination of DNA damage …

Cardiac, metabolic and growth response of early-stage chicken embryos to perturbations in yolk environment was investigated. Also, effects of parental hypoxia exposure on hypoxia resistance, thermal tolerance and body length of zebrafish larvae were investigated. In the first study, thyroxine, triiodothyronine and testosterone produced differential effects on heart rate and development rate of chicken embryos during the first 4 days of development. Triiodothyronine caused a dose-dependent increase in heart rate when applied at 40 or 70 hours of age, while thyroxine caused a dose-dependent increase in heart rate when applied at 40 hours only. Testosterone and propyl-thiouracil (deiodinase antagonist) did not have an effect on heart rate. Development rate was not changed by thyroxine, triiodothyronine, testosterone or propyl-thiouracil, which suggested that heart rate changes did not result from changes in embryo maturity. In the second study, chicken embryos exposed to yolks of different bird species during early-stage embryonic development showed changes in heart rate, mass-specific oxygen consumption and body mass that scaled with the egg mass, incubation period length, and yolk triiodothyronine and testosterone levels of the species from which yolk was derived. In the third study, this phenomenon was investigated between layer and broiler chickens. Heart rate, oxygen consumption …

Oxygen deprivation can be pathological for many organisms, including humans. Consequently, there are several biologically and economically relevant negative impacts associated with oxygen deprivation. Developing an understanding of which genes can influence survival of oxygen deprivation will enable the formulation of more effective policies and practices. In this dissertation, genes that influence adult anoxia survival in the model metazoan system, C. elegans, are identified and characterized. Insulin-like signaling, gonad function and gender have been shown to influence longevity and stress resistance in the soil nematode, C. elegans. Thus, either of these two processes or gender may influence anoxia survival. The hypothesis that insulin-like signaling alters anoxia survival in C. elegans is tested in Aim I. The hypotheses that gonad function or gender modulates anoxia survival are tested in Aim II. Insulin-like signaling affects anoxia survival in C. elegans. Reduction of insulin-like signaling through mutation of the insulin-like receptor, DAF-2, increases anoxia survival rates in a gpd-2/3 dependent manner. The glycolytic genes gpd-2/3 are necessary for wild-type response to anoxia, and sufficient for increasing anoxia survival through overexpression. Gonad function and gender both affect anoxia survival in C. elegans. A reduction of ovulation and oocyte maturation, as measured by oocyte flux, …

Fatty acid desaturase 2 (FAD2) enzymes are phosphatidylcholine desaturases occurring as integral membrane proteins in the endoplasmic reticulum membrane and convert monounsaturated oleic acid into polyunsaturated linoleic acid. The major objective of this research was to study the expression and function of two cotton FAD2 genes (the FAD2-3 and FAD2-4 genes) and their possible role in plant sensitivity to environmental stress, since plants may increase the polyunsaturated phospholipids in membranes under environmental stress events, such as low temperature and osmotic stress. Two FAD2 cDNA clones corresponding to the two FAD2 genes have been isolated from a cotton cDNA library, indicating both genes are truly expressed in cotton. Model yeast cells transformed with two cotton FAD2 genes were used to study the chilling sensitivity, ethanol tolerance, and growth rate of yeast cells. The expression patterns of the two FAD2 genes were analyzed by reverse transcription polymerase chain reactions (RT-PCR) and Western blot analyses in cotton plants under different treatment conditions. The coding regions of both FAD2 genes were inserted downstream from the CaMV 35S promoter in the pMDC gateway binary vector system. Five different FAD2/pMDC constructs were transformed into the Arabidopsis fad2 knockout mutant background, and multiple potential transgenic Arabidopsis plant …