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    Optimization and Characterization of the Intelligent Phagemid-Assembled Gene Expression (iPhAGE) Miniphagemid Gene Delivery Platform
    (University of Waterloo, 2026-07-17) Kosiak, Anna
    The Intelligent Phagemid-Assembled Gene Expression (iPhAGE) system is an M13 bacteriophage-derived, non-viral gene delivery platform. It produces miniaturized phagemid particles (miniphagemids): non-replicative, non-infectious particles that package circular single-stranded DNA (cssDNA) carrying a gene cassette with a gene of interest (GOI), such as a reporter or therapeutic gene. Unlike conventional viral vectors, miniphagemids package cssDNA cassettes devoid of bacterial backbone sequences, thereby reducing immunogenicity and addressing some regulatory concerns. Because miniphagemids cannot infect cells autonomously, traditional plaque-based quantification methods are inapplicable. Furthermore, key upstream production parameters, including genetic modifications to improve yield, growth media selection, and long-term storage stability, have not been systematically evaluated, yet are essential for the system's further development. This thesis addresses four key research gaps. First, ssDNA was identified as the most appropriate reference standard for quantifying the encapsidated genome in miniphagemids because it best reflects its single-stranded conformation and avoids topology-dependent amplification bias under the tested conditions. Second, in-house-developed SW8 helper plasmids achieve comparable yields to commercially available counterparts while exhibiting lower backbone contamination and superior packaging fidelity. Modification of the ribosome-binding site (RBS) in gV of the helper plasmid also improves yield; however, current findings suggest that this alteration alone is insufficient to enhance the output of pV-mediated ssDNA. Third, comparison of LB and TB growth media showed that, contrary to the expectation that nutrient-rich TB would increase titers, LB supported comparable or higher titers with lower backbone contamination and at a reduced cost under the conditions tested. Fourth, long-term stability was assessed over 120 days at various storage temperatures for the best-performing miniphagemids: DNA content was monitored by qPCR and NanoDrop; particle morphology and aggregation were evaluated by AFM and DLS; DNase protection assays assessed encapsidation integrity; and a luciferase-based transfection assay measured functional activity in the host mammalian cell line. Miniphagemids retained detectable DNA and functional transfection activity across all storage temperatures, though qPCR-based titers were best maintained at -20°C and 4°C, identifying these as optimal storage conditions. AFM and DLS revealed aggregation over time, and DNase protection assays confirmed that a second DNase treatment prior to quantification improves accuracy by removing residual non-encapsidated DNA. The SW8-EGF miniphagemid produced measurable luminescence without a transfection reagent after four months, consistent with receptor-mediated uptake. Taken together, these findings advance the characterization and process understanding of the iPhAGE system, providing a methodological framework for quantification, quality assessment, and production optimization of phagemid-derived particles.
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    Empirical evidence of the leverage effect in a stochastic volatility model: a realized volatility approach
    (University of Waterloo, 2010) Xu, Dinghai; Li, Yuying
    Increasing attention has been focused on the analysis of the realized volatility, which can be treated as a proxy for the true volatility. In this paper, we study the potential use of the realized volatility as a proxy in a stochastic volatility model estimation. We estimate the leveraged stochastic volatility model using the realized volatility computed from five popular methods across six sampling-frequency transaction data (from 1-min to 60-mi). Availability of the realized volatility allows us to estimate the model parameters via the MLE and thus avoids computational challenge in the high dimensional integration. Six stock indices are considered in the empirical investigation. We discover some consistent findings and interesting patterns from the empirical results. In general, the significant leverage effect is consistently detected at each sampling frequency. The volatility persistence becomes weaker at the lower sampling frequency. We also find that the consistent-scaling and "optimal"-weighted realized volatility method proposed by Hansen and Lunde (2005) provide relatively better performances compared to other methods considered.
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    Modeling asymmetric volatility clusters using copulas and high frequency data
    (University of Waterloo, 2010) Ning, Cathy; Xu, Dinghai; Wirjanto, Tony S.
    Volatility clustering is a well-known stylized feature of financial asset returns. In this paper, we investigate the asymmetric pattern of volatility clustering on both the stock and foreign exchange rate markets. To this end, we employ copula-based semi-parametric univariate time-series models that accommodate the clusters of both large and small volatilities in the analysis. Using daily realized volatilities of the individual company stocks, stock indices and foreign exchange rates constructed from high frequency data, we find that volatility clustering is strongly asymmetric in the sense that clusters of large volatilities tend to be much stronger than those of small volatilities. In addition, the asymmetric pattern of volatility clusters continues to be visible even when the clusters are allowed to be changing over time, and the volatility clusters themselves remain persistent even after forty days.
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    Evaluation of the impact of the increase in EI allowable earnings pilot project on working while on claim and job search behaviour in Canada
    (University of Waterloo, 2011-02) Lluis, Stephanie; McCall, Brian P.
    This paper examines the impact of the change in allowable earnings proposed in a pilot project (WWOC) of the Canadian Employment Insurance system implemented in December 2005 in some Canadian regions on working while on claim behaviour and on job search behaviour. The WWOC pilot is expected to increase the subsidy to low earning/part-time work. Search theory would predict that, all else equal, individuals would increase their intensity of search for these types of jobs. We find evidence that the WWOC pilot substantially increased the incidence and duration of work while on claim receiving full benefits and reduced the incidence and duration of working while on claim receiving no benefits for both men and women. We also find differences in the impact of the WWOC pilot on the job search behaviour of men. These results suggest that the WWOC pilot significantly encouraged working while on claim in low-paying jobs allowing receipt of full benefits. The WWOC pilot significantly reduced the number of hours looking for a job and reduced the likelihood of looking for only a full-time job (relative to looking for only a part-time job or either). These results are robust to the various robustness check analyses performed.
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    Uncertainty, task environment, and organization design: An empirical investigation
    (University of Waterloo, 2011-12) Ben-Ner, Avner; Kong, Fanmin; Lluis, Stephanie
    The paper addresses two broad research questions: 1. How do internal uncertainty associated with the task environment and external uncertainty arising from market volatility impact organization design? 2. What are the relationships among various elements of organization design: delegation of decision-making, incentives, monitoring, and internal labor market practices (promotion, training, employment security)? We expand on Prendergast (2002a), who challenged the conventional view of a tradeoff between risk and incentives, and build a single unified framework for answering our two research questions. Using a uniquely rich dataset that contains detailed information about the task environment of core employees and organization design at the individual, group and firms levels in 530 Minnesota firms in the mid 1990s, we first find support for Prendergast's key argument that internal uncertainty (over which employees have control) affects directly the allocation of decision-making and only indirectly incentives (via allocation of decision-making). This confirms similar findings by Foss and Laursen (2005), DeVaro and Kurthulus (2010) and Shi (2011). We also find that internal uncertainty has much impact on organization design through the choice of delegation of decision-making at the employee level, less so at the group level, and very little at the firm level, whereas external (market) uncertainty has little effect on organization design, especially at the individual and group level. Decision-making, monitoring, various internal labor market practices and incentives are strongly related to each other through substitution and complementarity.