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The sustainability of the beef industry has become a point of national interest, particularly the investment of land and water resources. Our objective was to estimate how much land and irrigation water are required to maintain a simulated Angus cow-calf operation in the North Central Great Plains (NCGP) for an average year. A stochastic model was used, which enabled consideration of biological variation. The model computed 100 iterations of a 24-year timeframe (1995–2018). The simulated herd had 100 breeding females with replacement heifers being retained annually. The nutrients required to maintain a body condition score 5 for each individual animal, adjusting for temperature and physiological state, were calculated. A stocking rate of 3.3 hectares (ha) per cow-calf pair and mature cow weight of 600 kg was set, which is representative of the NCGP. Replacement heifers were assumed to be 65% of mature cow weight and allotted 1.22 ha. Bred heifers were assumed to be 85% of mature cow weight and allotted 1.81 ha. The herd was assumed to be grazing from May 1 to October 31. A supplemented ration of 60% alfalfa and 40% corn was provided if an individual’s nutritional needs were not met. Animals were assumed to be delivered a base ration from November 1 to April 30, which consisted of 73% alfalfa, 19% wheat straw, and 8% corn. The amount of irrigation necessary to grow feed was determined by estimating evapotranspiration of each crop then subtracting the amount of precipitation during the growing season. Average crop yield was determined using county level data from the UDSA NASS to estimate how much land would be needed for feed production. Sustaining a 100 head cow-calf herd in the NCGP for an average year requires 103.5 million liters for irrigation, 1288.5 ha for crop production, 357 ha grazing land.

Leishmaniasis is caused by the flagellated protozoan Leishmania, and is a neglected tropical disease (NTD), as defined by the World Health Organisation (WHO). Bisulphite conversion technology converts all genomic material to a simplified form during the lysis step of the nucleic acid extraction process, and increases the efficiency of multiplex quantitative polymerase chain reaction (qPCR) reactions. Through utilization of qPCR real-time probes, in conjunction with bisulphite conversion, a new duplex assay targeting the 18S rDNA gene region was designed to detect all Leishmania species. The assay was validated against previously extracted DNA, from seven quantitated DNA and cell standards for pan-Leishmania analytical sensitivity data, and 67 cutaneous clinical samples for cutaneous clinical sensitivity data. Specificity was evaluated by testing 76 negative clinical samples and 43 bacterial, viral, protozoan and fungal species. The assay was also trialed in a side-by-side experiment against a conventional PCR (cPCR), based on the Internal transcribed spacer region 1 (ITS1 region). Ninety-seven percent of specimens from patients that previously tested positive for Leishmania were positive for Leishmania spp. with the bisulphite conversion assay, and a limit of detection (LOD) of 10 copies per PCR was achieved, while the LOD of the ITS1 methodology was 10 cells/1000 genomic copies per PCR. This method of rapid, accurate and simple detection of Leishmania can lead to improved diagnosis, treatment and public health outcomes.

The aim of this thesis is the development and implementation of a controlled testing platform for the Robust Intelligent Sensing and Controls (RISC) Lab at Utah State University (USU). This will be an open source adaptable expandable robotics platform usable for both education and research. This differs from the many other platforms developed in that the entire platform software will be made open source. This open source software will encourage collaboration among other universities and enable researchers to essentially pick up where others have left off without the necessity of replicating months or even years of work. The expected results of this research will create a foundation for diverse robotics investigation at USU as well as enable attempts at novel methods of control, estimation and optimization. This will also contribute a complete software testbed setup to the already vibrant robotics open source research community. This thesis first outlines the platform setup and novel developments therein. The second stage provides an example of how this has been used in education, providing an example curriculum implementing modern control techniques. The third section provides some exploratory research in trajectory control and state estimation of the tip of an inverted pendulum atop a small unmanned aerial vehicle as well as bearing-only cooperative localization experimentation. Finally, a conclusion and future work is discussed.

Canonical models of rational choice fail to account for many forms of motivated adaptive behaviors, specifically in domains such as food selections. To describe behavior in such emotion- and reward-laden scenarios, researchers have proposed dual-process models that posit competition between a slower, analytic faculty and a fast, impulsive, emotional faculty. In this paper, we examine the assumptions and limitations of these approaches to modeling motivated choice. We argue that models of this form, though intuitively attractive, are biologically implausible. We describe an approach to motivated choice based on sequential sampling process models that can form a solid theoretical bridge between what is known about brain function and environmental influences upon choice. We further suggest that the complex and dynamic relationships between biology, behavior, and environment affecting choice at the individual level must inform aggregate models of consumer choice. Models using agent-based complex systems may further provide a principled way to relate individual and aggregate consumer choices to the aggregate choices made by businesses and social institutions. We coin the term "brain-to-society systems" choice model for this broad integrative approach.

Inhibin is composed of an alpha- and a beta-subunit. Transgenic studies assigned a tumor-suppressive role to the inhibin alpha-subunit, and in human prostate cancer inhibin alpha-subunit gene expression was down-regulated. This study examined the inhibin alpha-subunit gene promoter and gene locus to determine whether promoter hypermethylation or LOH occurred in DNA from prostate cancer. The 5'-untranslated region of the human inhibin alpha-subunit gene was sequenced and shown to be highly homologous to the bovine, rat, and mouse inhibin alpha-subunit promoter sequences. A 135-bp region of the human promoter sequence that continued a cluster of CpG sites was analyzed for hypermethylation. Significant (P0.001) hypermethylation of the inhibin alpha-subunit gene promoter occurred in DNA from Gleason pattern 3, 4, and 5 carcinomas compared with nonmalignant tissue samples. A subset of the carcinomas with a cribriform pattern were unmethylated. LOH at 2q32-36, the chromosomal region harboring the inhibin alpha-subunit gene, was observed in 42% of prostate carcinomas. These data provide the first demonstration that promoter hypermethylation and LOH are associated with the inhibin alpha-subunit gene and gene locus in prostate cancer.

Glutathione-S-Transferases (GSTs) comprise a family of isoenzymes that provide protection to mammalian cells against electrophilic metabolites of carcinogens and reactive oxygen species. Previous studies have shown that the CpG-rich promoter region of the pi-class gene GSTP1 is methylated at single restriction sites in the majority of prostate cancers. In order to understand the nature of abnormal methylation of the GSTP1 gene in prostate cancer we undertook a detailed analysis of methylation at 131 CpG sites spanning the promoter and body of the gene. Our results show that DNA methylation is not confined to specific CpG sites in the promoter region of the GSTP1 gene but is extensive throughout the CpG island in prostate cancer cells. Furthermore we found that both alleles are abnormally methylated in this region. In normal prostate tissue, the entire CpG island was unmethylated, but extensive methylation was found outside the island in the body of the gene. Loss of GSTP1 expression correlated with DNA methylation of the CpG island in both prostate cancer cell lines and cancer tissues whereas methylation outside the CpG island in normal prostate tissue appeared to have no effect on gene expression.

Methylation analysis of individual cytosines in genomic DNA can be determined quantitatively by bisulphite treatment and PCR amplification of the target DNA sequence, followed by restriction enzyme digestion or sequencing. Methylated and unmethylated molecules, however, have different sequences after bisulphite conversion. For some sequences this can result in bias during the PCR amplification leading to an inaccurate estimate of methylation. PCR bias is sequence dependent and often strand-specific. This study presents a simple method for detection and measurement of PCR bias for any set of primers, and investigates parameters for overcoming PCR bias.

The modulation of DNA-protein interactions by methylation of protein-binding sites in DNA and the occurrence in genomic imprinting, X chromosome inactivation, and fragile X syndrome of different methylation patterns in DNA of different chromosomal origin have underlined the need to establish methylation patterns in individual strands of particular genomic sequences. We report a genomic sequencing method that provides positive identification of 5-methylcytosine residues and yields strand-specific sequences of individual molecules in genomic DNA. The method utilizes bisulfite-induced modification of genomic DNA, under conditions whereby cytosine is converted to uracil, but 5-methylcytosine remains nonreactive. The sequence under investigation is then amplified by PCR with two sets of strand-specific primers to yield a pair of fragments, one from each strand, in which all uracil and thymine residues have been amplified as thymine and only 5-methylcytosine residues have been amplified as cytosine. The PCR products can be sequenced directly to provide a strand-specific average sequence for the population of molecules or can be cloned and sequenced to provide methylation maps of single DNA molecules. We tested the method by defining the methylation status within single DNA strands of two closely spaced CpG dinucleotides in the promoter of the human kininogen gene. During the analysis, we encountered in sperm DNA an unusual methylation pattern, which suggests that the high methylation level of single-copy sequences in sperm may be locally modulated by binding of protein factors in germ-line cells.

Culture of mesenchymal stem cells (MSCs) under conditions promoting proliferation and differentiation, while supporting genomic and epigenetic stability, is essential for therapeutic use. We report here the extent of genome-wide DNA gains and losses and of DNA methylation instability on 170 cancer-related promoters in bone marrow (BM) MSCs during culture to late passage in medium containing fetal bovine serum (FBS) or autologous serum (AS). Comparative genomic hybridization indicates that expansion of BMMSCs elicits primarily telomeric deletions in a subpopulation of cells, the extent of which varies between donors. However, late passage cultures in AS consistently display normal DNA copy numbers. Combined bisulfite restriction analysis and bisulfite sequencing show that although DNA methylation states are overall stable in culture, AS exhibits stronger propensity than FBS to maintain unmethylated states. Comparison of DNA methylation in BMMSCs with freshly isolated and cultured adipose stem cells (ASCs) also reveals that most genes unmethylated in both BMMSCs and ASCs in early passage are also unmethylated in uncultured ASCs. We conclude that (i) BMMSCs expanded in AS or FBS may display localized genetic alterations, (ii) AS tends to generate more consistent genomic backgrounds and DNA methylation patterns, and (iii) the unmethylated state of uncultured MSCs is more likely to be maintained in culture than the methylated state.

This research was undertaken to investigate the effects of solid modeling and visualization on technical problem solving. The participants were 47 students enrolled in solid modeling classes at Southeast Missouri State University. The control and experimental groups consisted of 23 and 24 randomly assigned students respectively. This study was a posttest only design that used logistic regression to analyze the results. Both groups were required to take the Purdue Spatial-Visualization Test/Visualization of Rotations (PSVT/TR). Participants in the control group used only sketching to design their solutions while participants in the experimental group used parametric solid modeling software to design their solutions. All participants then constructed prototypes of their designs. The prototype was evaluated to determine if it successfully met the design specifications. The findings revealed that visualization was a significant predictor of technical problem solving as defined by successful prototype construction (p=.021). There was no significant difference between the sketching and solid modeling design methods used for technical problem solving (p=.752). The interaction between the method of design, solid modeling or sketching, was analyzed to determine if using solid modeling would offset low visualization scores It was found that the interaction was not significant (p=.393). Ph. D.

What defines the boundaries between methylated and unmethylated domains in the genome is unclear. In this study we used bisulfite genomic sequencing to map the boundaries of methylation that flank the 5′- and 3′-ends of the CpG island spanning the promoter region of the glutathione S-transferase (GSTP1) gene. We show that GSTP1 is expressed in a wide range of tissues including brain, lung, skeletal muscle, spleen, pancreas, bone marrow, prostate, heart, and blood and that this expression is associated with the CpG island being unmethylated. In these normal tissues a marked boundary was found to separate the methylated and unmethylated regions of the gene at the 5′-flank of the CpG island, and this boundary correlated with an (ATAAA)19–24 repeated sequence. In contrast, the 3′-end of the CpG island was not marked by a sharp transition in methylation but by a gradual change in methylation density over about 500 base pairs. In normal tissue the sequences on either side of the 5′-boundary appear to lie in separate domains in which CpG methylation is independently controlled. These separate methylation domains are lost in all prostate cancer whereGSTP1 expression is silenced and methylation extends throughout the island and spans across both the 5′- and 3′-boundary regions.

Summary: The technique of representation difference analysis PCR has been applied to find genes specific to Mycobacterium avium subsp. paratuberculosis. This generated a 671 bp fragment which was used to isolate a larger genetic element found in the enteric pathogens M. avium subsp. paratuberculosis and M. avium subsp. silvaticum but which was absent from the very closely related and relatively benign M. avium subsp. avium. This element, designated GS, is greater than 6·5 kbp in length and has a G+C content 9 mol% lower than other genes from this species. There is a previously uncharacterized insertion sequence associated with one end. The GS element encodes five ORFs in M. avium subsp. paratuberculosis and M. avium subsp. silvaticum, all of which have counterparts encoded in Mycobacterium tuberculosis. Database searches revealed homologues for these ORFs in a number of bacterial species, predominantly Gram-negative organisms, including a number of enteric pathogens. These homologous genes encode functions related to LPS or extracellular polysaccharide biosynthesis. This element has a number of features in common with pathogenicity islands such as its low G+C content, an association with a putative insertion sequence and a grouping of genes of related function with a possible link to virulence. No direct link to pathogenicity has been shown but GS may belong to a group of related ‘genetic islands’ and represents the first such element to be identified in mycobacteria.

The Mycobacterium avium subsp. paratuberculosis (formerly Mycobacterium paratuberculosis) atypical insertion sequence, IS900, encodes a novel gene on the complementary strand to the putative transposase, p43. This gene requires a promoter, ribosome binding site (RBS) and termination codon to be acquired upon insertion into the M. avium subsp. paratuberculosis genome and hence is designated the hed (host expression-dependent) gene of IS900. Analysis of IS900 insertion sites suggests that this element targets translation initiation signals in M. avium subsp. paratuberculosis, specifically inserting between the RBS and start codon of a putative gene sequence. This aligns the hed initiation codon adjacent to a functional RBS and possibly downstream of an active promoter, driving expression of Hed protein. We have confirmed this unique targeting process by detecting expression of hed in M. avium subsp. paratuberculosis at the level of transcription by reverse transcription-PCR. Further, two Hed-specific antibodies detected Hed translation products in Western blots of protein extracts from M. avium subsp. paratuberculosis. A recombinant form of Hed expressed and purified from Escherichia coli will facilitate studies of IS900 transposition and will also be assessed as a diagnostic antigen for M. avium subsp. paratuberculosis disease. Implications of IS900 insertion in M. avium subsp. paratuberculosis pathogenicity are discussed.