Thesis (M.A. in agribusiness)--Washington State University. / Includes bibliographical references.

This thesis analyses the development of safety regulation for the intentional release of genetically modified organisms (GMOs) between 19 89-92, especially in Britain in its European context, and by contrast to the USA. The thesis emphasizes the practical dilemmas of GMO regulation in accommodating uncertainties about public unease and environmental harm. It serves as a case study of safety regulation as a constructed rationality, of national regulatory styles, and of environmental precaution. In anticipating hazards prior to evidence of harm, GMO regulation had a contested 'rational' basis. Regulators encountered disputes in defining the risk problem, in establishing risk-management institutions, and in reducing scientific uncertainty about potential harm. Insofar as GMO regulation had a precautionary content, it undermined the 'rational' stereotype of risk-assessment steps. Both the precautionary potential and its limits derived from the project of overcoming obstacles to a biotechnology market. This meant symbolically normalizing GMOs as benign products, while specifying testable ecological uncertainties rooted in some naturalistic analogy. Technical 'risk' abstracted potential harm from issues of socioagronomic control which underlay the earlier environmental controversy. The thesis argues for recasting theoretical models of safety regulation as a 'technical' or 'procedural' rationality. GMO regulation contained poles of tension which such theoretical models attribute to antagonistic rationalities. Broadly speaking, the regulatory system was managing an internal contradiction between social legitimacy and commercialization. The difficulties of GMO regulation arose from its implicit role in legitimizing biotechnology, by default of any democratic procedure for adjudicating a contentious technoscientific development.

Thesis (Ph. D.)--University of Toronto, 2000. / Includes bibliographical references (p. 413-445). Also available on the World Wide Web.

No description available.

A luminescence based marker system was developed for detection of genetically modified Pseudomonas fluorescens and E. coli. During batch growth in liquid culture, luminescence measured by luminometry was directly proportional to biomass concentration and enabled detection of 104 - 106 cells ml-1 of P. fluorescens and 101 cells ml-1 of E. coli, in actively growing cultures. Following inoculation into soil, detection levels were reduced ten-fold. After the subsequent utilisation of available nutrients, activity and luminescence decreased and luminometry then provided a direct, non-extractive means of measuring population activity of lux-marked inocula. Potential luminescence, measured as luminescence following amendment with nutrients, enabled assessment of the rate of reactivation of the lux-marked inocula and quantification of the size of the activatable population. Both these techniques, and traditional techniques, were used to investigate the survival of P. fluorescens and E. coli in soil microcosms. The effect of matric potential and indigenous organisms on luminescence and on survival of P. fluorescens was assessed. Matric potential significantly decreased the activity of both introduced and indigenous populations, but the indigenous population also significantly decreased the activity and biomass concentration of the introduced P. fluorescens population. Use of luminometry as a non-extractive measure of biomass concentration provided qualitative correlation with viable cell concentration, suggesting its potential for rapid enumeration of marked inocula. Reactivation of cells at increased matric stress was decreased, but use of high substrate:cell ratios at -30 kPa produced higher levels of luminescence and may, therefore, improve the use of luminometry as an estimate of biomass.

Methods currently available for the detection and enumeration of genetically modified micro-organisms in the environment include culturing methods, direct microscopic detection and nucleic hybridization techniques. The aim of this project was to develop luminescence as a molecular based-marker system in Pseudomonas fluorescens. The lux genes, originally isolated from Vibrio fischeri, were introduced into Ps. fluorescens on plasmid vectors and on the chromosome. The efficiency of these two strategies for the detection of Ps. fluorescens in soil was assessed. Luminometry was used to estimate biomass concentration during growth. The sensitivity of luminescence detection was greater for the plasmid marked Ps. fluorescens in both liquid culture and soil, however, cellular light output was less closely linked to biomass concentration. Enumeration of cells by luminometry was only possible for growing cells as light output is correlated with microbial activity. The lux chromosomal marker was stable in liquid culture for at least 200 generations and in soil for up to 135 days. The plasmid borne lux genes had a half-life of 20 generations in liquid culture. After inoculation in sterile soil, plasmid loss was only observed during cellular growth. The frequency of transfer of the lux genes from Ps. fluorescens, by conjugation and transformation, was assessed in liquid culture. Mobilisation of these genes by three self transmissible plasmids was negligible due to the instability of these vectors in this host. Transformation of Ps. stutzeri with lux genes, by cell contact, was at frequencies below levels of detection. Luminescence provided a valuable marker for tracking pseudomonads in soil. Detection of marked strains by luminometry provided a sensitive, rapid and non-extractive technique for enumeration of growing cells and measurement of microbial activity. As the chromosomally encoded lux genes were stable, regardless of growth conditions, and emitted sufficient levels of light to enable visual enumeration of colonies by eye, this was considered the best system for long term risk assessment studies.

The long-term implications of deregulation of glucose metabolism, manifesting itself as diabetes is significant. Whilst treatments for the disease are progressing, often they require multiple daily insulin injections. This may be difficult for diabetics in poorer nations. This thesis examines the possibility of utilising keratinocytes and melanocytes transfected with either normal or furin-modified proinsulin constructs, to process and secrete mature insulin via secreted protein pathways. The advantage of this approach is that it may allow normalisation of basal glucose levels, which in turn may prevent or delay the onset of many of the complications associated with diabetes. The results confirmed that keratinocytes and melanocytes are able to process and secrete mature insulin from a furin-modified proinsulin construct, thus validating this approach and paving the way for skin grafts of genetically modified cells to be used as a possible treatment for diabetes. Whilst the results regarding the ability of these cells to process a normal proinsulin construct are inconclusive, they highlight areas of interest that could help to resolve this issue / Master of Science (Hons.)

Thesis (B.A.) Summa Cum Laude --Butler University, 2009. / Includes bibliographical references (leaves 44-47).

The aim of this work was to develop and test a mathematical model for DNA transfer by conjugation which considered spatial separation of cells on surfaces. Model parameters include the initial donor and recipient numbers, maximum specific population growth rate, microcolony radial extension rate, maximum cell yield, maximum incubation time, standard deviation of the mean intercellular distance (σ), cellular area and area available for colonization. Sensitivity analysis indicated that growth areas and σ had the greatest effects on conjugation. The model was extended to consider the presence of a strain that did not conjugate and its effect on plasmid transfer. Filter matings with <I>Pseudomonas fluorescens</I> MON787 RP4 as the plasmid donor and <I>P. fluorescens</I> MON787 R <I>lux</I> as recipient were used to test the model. Model predictions were generally accurate but transconjugants were consistently underestimated. This was attributed to intercellular distances not following a strict Gaussian distribution. Nevertheless, predicted and experimental data were qualitatively similar, which increased confidence in the validity of the mechanisms proposed. Conjugation occurred over a wide range of cell densities, donor to recipient ratios, nutrient levels and incubation temperatures. Starved cells retained the capacity to conjugate, but plasmid transfer frequency was higher in the presence of nutrient. Above a minimum level, conjugation ability was not enhanced by nutrients. Temperature affected conjugation ability, the optimum being 20°C - 30°C. The presence of a nonconjugative strain decreased conjugation, by leading to earlier nutrient exhaustion and growth arrest which limited meetings between donor and recipient microcolonies. The model and the experimental system demonstrate the importance of spatial effects on conjugation. Description and prediction of gene transfer in natural environments will require models of greater complexity, and more sophisticated experimental testing, but this study provides a basis for theoretical descriptions of gene transfer in heterogeneous natural environments, such as soil and biofilms on solid surfaces in aquatic environments.

No description available.