published_or_final_version / Medicine / Doctoral / Doctor of Philosophy

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University of Macau / Institute of Chinese Medical Sciences

by Lam Ying Hoo. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1994. / Includes bibliographical references (leaves 145-149). / Acknowledgements --- p.i / Abstract --- p.ii / Contents --- p.iv / Abbreviations --- p.x / Chapter Chapter 1 --- Introduction / Chapter 1.1. --- Trichosanthin --- p.1 / Chapter 1.2. --- Anti-plant viral and fungal properties of RIPs --- p.3 / Chapter 1.3. --- Agrobacterium-mediated transformation --- p.5 / Chapter 1.3.1. --- Ti (tumor inducing) plasmid --- p.6 / Chapter 1.3.2. --- Role of vir proteins in T-DNA transfer --- p.6 / Chapter 1.3.3. --- Integration of T-DNA into plant genome --- p.11 / Chapter 1.3.4. --- Use of Agrobacterium plasmid as transformation vectors --- p.13 / Chapter 1.4. --- Objective and strategy of producing transgenic plants that express TCS --- p.15 / Chapter Chapter 2 --- Materials and Methods / Chapter 2.1. --- Bacterial Strains used --- p.19 / Chapter 2.2. --- General Techniques --- p.20 / Chapter 2.2.1. --- Growth of bacterial strains --- p.20 / Chapter 2.2.2. --- Restriction Enzyme Digestion of DNA --- p.21 / Chapter 2.2.3. --- Agarose Gel Electrophoresis of DNA --- p.21 / Chapter 2.2.4. --- Purification of DNA fragments from Agarose Gel using GeneClean II® ( BIO 101 Inc.) kit --- p.22 / Chapter 2.2.5. --- Purification of DNA fragments by Phenol/Chloroform Extraction --- p.23 / Chapter 2.2.6. --- Ligation of DNA fragments --- p.24 / Chapter 2.2.7. --- Preparation and Transformation of Escherichia coli Competent Cells --- p.24 / Chapter 2.2.8. --- Minipreparation of Plasmid DNA --- p.26 / Chapter 2.2.9. --- Preparation of Plasmid DNA using Magic´ёØ Minipreps DNA Purification kit from Promega --- p.27 / Chapter 2.2.10. --- Preparation of Plasmid DNA using Qiagen-pack 100 Cartridge --- p.29 / Chapter 2.2.11. --- SDS-Polyacrylamide Gel Electrophoresis (SDS-PAGE) --- p.30 / Chapter 2.2.12. --- Western Blot detection of TCS --- p.33 / Chapter 2.2.13. --- Polymerase Chain Reaction (PCR) --- p.34 / Chapter 2.3. --- Construction of Plant Transformation Vectors --- p.36 / Chapter 2.3.1. --- Construction of pSLJ58210 --- p.36 / Chapter 2.3.2. --- Construction of pSLJ TCS1 and pSLJ TCS2 --- p.38 / Chapter 2.3.3. --- Conjugation of pSLJ TCS1 and pSLJ TCS2 into A. tumefaciens by Triparental Mating --- p.41 / Chapter 2.4. --- Transformation of Tobacco Leaf Explants by Agrobacterium tumefaciens --- p.43 / Chapter 2.4.1. --- Growth of A. tumefaciens LBA4404 (pSLJ TCS1) --- p.43 / Chapter 2.4.2. --- Surface Sterilization of tobacco leaves --- p.43 / Chapter 2.4.3. --- Inoculation of tobacco leaf explants with A. tumefaciens LBA4404 (pSLJ TCS1) --- p.44 / Chapter 2.4.4. --- Regeneration of shoots from Transformed explants --- p.45 / Chapter 2.4.5. --- Rooting of Transformed shoots --- p.45 / Chapter 2.4.6. --- Re-establishment of cultured Plantlets in soil --- p.45 / Chapter 2.5. --- Analysis of the Regenerated Transgenic Tobacco --- p.46 / Chapter 2.5.1. --- Isolation of plant leaf protein --- p.46 / Chapter 2.5.2. --- SDS-PAGE and Western blot detection of TCS --- p.48 / Chapter 2.5.3. --- Anti-viral assay of Transgenic tobacco against TuMV --- p.48 / Chapter 2.6. --- Bioassay of Inhibitory activity of TCS protein against TuMV --- p.49 / Chapter 2.6.1. --- Preparation of biologically active TCS protein --- p.49 / Chapter 2.6.2. --- Purification of TuMV from infected plant leaves --- p.51 / Chapter 2.6.3. --- Mechanical Inoculation of virus onto host plant --- p.53 / Chapter 2.6.4. --- Anti-viral assay on Local Lesion host --- p.54 / Chapter 2.6.5. --- Anti-viral assay on Systemic host --- p.55 / Chapter 2.7 --- Establishment of the plant culture medium for efficient Regeneration from tissue explants of Brassica parachinensis --- p.56 / Chapter 2.7.1. --- Preparation and Sterilization of culture medium --- p.57 / Chapter 2.7.2. --- Preparation of Sterile seedlings of B. parachinensis --- p.57 / Chapter 2.7.3. --- Regeneration from Cotyledon petiole and Hypocotyl segment explants --- p.58 / Chapter 2.7.4. --- Regeneration from Internode stem segment explants of shoot culture --- p.60 / Chapter 2.8. --- Reagents and Buffers --- p.61 / Chapter 2.8.1. --- Media for Bacterial culture --- p.61 / Chapter 2.8.2. --- Media for Plant tissue culture --- p.64 / Chapter 2.8.3. --- Restriction Enzymes --- p.66 / Chapter 2.8.4. --- Buffers for Agarose Gel Electrophoresis --- p.66 / Chapter 2.8.5. --- DNA ligation Buffer --- p.67 / Chapter 2.8.6. --- Reagents for preparation of E.coli competent cells --- p.67 / Chapter 2.8.7. --- Reagents for preparation of Plasmid DNA --- p.68 / Chapter 2.8.8. --- Reagents for Qiagen-pack 100 Cartridge --- p.69 / Chapter 2.8.9. --- Reagents for SDS-PAGE --- p.70 / Chapter 2.8.10. --- Reagents for Western Blotting --- p.71 / Chapter Chapter 3 --- Construction of Plant Transformation Vectors / Chapter 3.1. --- Introduction --- p.73 / Chapter 3.2. --- Results --- p.74 / Chapter 3.2.1. --- Construction of pSLJ58210 --- p.74 / Chapter 3.2.2. --- Construction of the recombinant binary vectors pSLJ TCSl and pSLJ TCS --- p.78 / Chapter 3.2.3. --- Conjugation ofpSLJ TCS 1 and pSLJ TCS 2 into Agrobacterium tumefaciens via Triparental Mating --- p.82 / Chapter 3.3. --- Discussion --- p.90 / Chapter Chapter 4 --- Transformation of Tobacco Leaf Explants by Agrobacterium tumefaciens / Chapter 4.1. --- Introduction --- p.94 / Chapter 4.2. --- Results --- p.95 / Chapter 4.2.1. --- Regeneration of leaf explants after transformation --- p.95 / Chapter 4.2.2. --- The level of expression of TCS in transgenic tobacco leaf --- p.100 / Chapter 4.3. --- Discussion --- p.104 / Chapter 4.3.1. --- Regeneration of transgenic tobacco plants --- p.104 / Chapter 4.3.2. --- Expression of TCS in transgenic tobacco plants --- p.108 / Chapter Chapter 5 --- Two approaches to study the Inhibitory effect of TCS on TuMV / Chapter 5.1. --- Introduction --- p.112 / Chapter 5.2. --- Results --- p.113 / Chapter 5.2.1. --- Expression and purification of recombinant TCS --- p.113 / Chapter 5.2.2. --- Purification of TuMV --- p.119 / Chapter 5.2.3. --- Anti-viral assay on local lesion host --- p.119 / Chapter 5.2.4. --- Anti-viral assay on Systemic host --- p.124 / Chapter 5.2.5. --- Anti-viral assay of Transgenic tobacco against TuMV --- p.126 / Chapter 5.3. --- Discussion --- p.129 / Chapter Chapter 6 --- Establishment of plant culture conditions for efficient shoot regeneration from tissue explants of B.parachinensis / Chapter 6.1. --- Introduction --- p.133 / Chapter 6.2. --- Results --- p.133 / Chapter 6.3. --- Discussion --- p.137 / Chapter Chapter 7 --- Conclusion / Appendix / Chapter A.1. --- Size of molecular weight markers --- p.143 / Chapter A.2. --- References --- p.145

Thesis (MMedVet (Anaes) (Companion Animal Clinical Studies, Veterinary Science))--University of Pretoria, 2009. / Includes bibliographical references. Also available in print format.

published_or_final_version / Medicine / Master / Doctor of Medicine

Tuberculosis (TB) is the leading cause of death in individuals infected with human immunodeficiency virus (HIV) in several African countries, including South Africa. HIV-positive individuals do not have the immune system resources to keep TB in check and are as much as 30 times more likely to develop active TB than people who are HIV-negative. Many people infected with HIV develop TB as the first manifestation of AIDS and TB accelerates disease progression in HIV-positive individuals. HIV and TB pathogenesis are thus inextricably intertwined so that it is necessary for medical practitioners to have an understanding of the dynamics and treatment of HIV-TB coinfection. At present the question remains as to whether the best time for coinfected individuals to start antiretroviral treatment for HIV is at the beginning, the peak, or after the completion of the TB treatment phase. This dissertation was undertaken with the aim of obtaining some clarity on this question by creating a mathematical model of HIV-TB coinfection and its treatment. This needs an understanding of the biological interactions; therefore the dissertation begins with a discussion of the biological mechanisms of HIV, the human immune system, TB and the drug therapies for each disease. Thereafter a brief introduction to mathematical modelling reviews basic HIV models, which are then modified to include HIV drug therapy. Analyses and simulations of these models were carried out, which yielded some insights into the dynamics of HIV and HIV therapy. Finally HIV-TB coinfection is introduced by reviewing a previously developed model. Based on all the models reviewed, a model for coinfection is developed which includes treatment for HIV and TB. Numerical simulations suggest that, if HIV disease progression is at an advanced stage of the immune system collapsing towards AIDS, with low T-cell count and high viral load, it is necessary to treat for both diseases simultaneously to ensure a positive survival prognosis for the coinfected individual. However, if disease progression is in the early stages of AIDS, with T-cell count and viral load beginning to display signs of the immune system collapse but still at reasonable levels relative to advanced stages, it need not be necessary to treat both diseases simultaneously. TB can be treated first, and upon completion HIV treatment can be initiated thus sparing the coinfected individual from the compounded side-effects and drug-drug interactions which usually result from simultaneous treatment. / Thesis (M.Sc.)-University of KwaZulu-Natal, 2007.

L-carnitine is an endogenous compound that has important roles in fatty acid oxidation. Patients with end-stage renal disease (ESRD) who are undergoing haemodialysis may develop a secondary L-carnitine deficiency. Following oral administration of L-carnitine, enterobacteria generate ??-butyrobetaine and trimethylamine with the latter substance extensively N-oxygenated in the liver, to form trimethylamine-N-oxide. Given that patients with ESRD have qualitatively different and higher bacterial populations in the small intestine as compared with healthy subjects, increased formation of trimethylamine and accumulation of trimethylamine-N-oxide would be expected. The clinical significance of these amines is related to their potential to form the carcinogen N-nitrosodimethylamine, contribution to neurological toxicity and "uraemic breath". / The pharmacokinetics of oral L-carnitine display clear non-linearity above a dose of 0.5 g three times a day with an associated increase in plasma concentrations of trimethylamine and trimethylamine-N-oxide. Oral administration of L-carnitine to patients with ESRD undergoing haemodialysis increased plasma concentrations of this substance to levels seen in individuals with normal kidney function and evidence was provided for the accumulation of trimethylamine-N-oxide. / Thesis (PhD)--University of South Australia, 2006.