by Chan, Yuk Wah. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2009. / Includes bibliographical references (leaves 166-175). / Abstracts in English and Chinese. / ACKNOWLEDGEMENTS --- p.iii / ABSTRACT --- p.v / 摘要 --- p.vii / LIST OF CONTENTS --- p.viii / LIST OF FIGURES --- p.xiv / LIST OF TABLES --- p.xvii / LIST OF ABBREVIATIONS --- p.xviii / Chapter CHAPTER 1. --- GENERAL INTRODUCTION --- p.1 / Chapter CHAPTER 2. --- LITERATURE REVIEW --- p.5 / Chapter 2.1 --- Vitamin A --- p.5 / Chapter 2.1.1 --- General and properties --- p.5 / Chapter 2.1.2 --- Biological importance of vitamin A --- p.6 / Chapter 2.1.3 --- Deficiency symptoms --- p.9 / Chapter 2.1.4 --- Dietary source of vitamin A --- p.10 / Chapter 2.1.5 --- Metabolism of dietary vitamin A and provitamin A in human --- p.12 / Chapter 2.1.5.1 --- Digestion and absorption --- p.12 / Chapter 2.1.5.2 --- Bioconversion --- p.12 / Chapter 2.1.5.2.1 --- "β, β-carotene-15,15'-monooxygenase (BCMO)" --- p.13 / Chapter 2.1.5.3 --- "Transport, uptake and storage" --- p.15 / Chapter 2.2 --- Vitamin A deficiency (VAD) --- p.19 / Chapter 2.2.1 --- Present situation --- p.19 / Chapter 2.2.2 --- Global efforts in dealing with VAD --- p.21 / Chapter 2.2.2.1 --- Vitamin A supplementation --- p.21 / Chapter 2.2.2.2 --- Food fortification --- p.22 / Chapter 2.2.2.3 --- Biofortification --- p.23 / Chapter 2.2.2.3.1 --- Conventional selective breeding --- p.23 / Chapter 2.2.2.3.2 --- Biosynthesis of provitamin A in plants --- p.25 / Chapter 2.2.2.3.3 --- Carotenoids enhancement in major plants and food crops --- p.31 / Chapter 2.3 --- Inherent problems of the present carotenoid enhancement --- p.34 / Chapter 2.3.1 --- Recommended Dietary Amount of vitamin A --- p.34 / Chapter 2.3.2 --- Factors affecting the bioefficacy of provitamin A in human body --- p.35 / Chapter 2.3.2.1 --- Bioavailability --- p.36 / Chapter 2.3.2.2 --- Bioconvertibility --- p.38 / Chapter 2.3.2.3 --- Health and nutrition status --- p.39 / Chapter 2.4 --- Previous study in our lab --- p.41 / Chapter 2.4.1 --- Overexpression of rice PSY1 --- p.41 / Chapter 2.4.2 --- Introduction of carotenoid genes and BCMOs into rice --- p.44 / Chapter 2.5 --- Overview of the project --- p.50 / Chapter CHAPTER 3. --- MATERIALS AND METHODS --- p.52 / Chapter 3.1 --- Chemicals --- p.52 / Chapter 3.2 --- Bacterial strains in regular cloning --- p.52 / Chapter 3.3 --- BCMO genes and carotenogenic genes --- p.53 / Chapter 3.4 --- Expression of BCMOs in bacterial system --- p.54 / Chapter 3.4.1 --- lac promoter system --- p.54 / Chapter 3.4.2 --- pBAD-TOPO® system --- p.56 / Chapter 3.5 --- Construction of gene cassettes for plant transformation --- p.58 / Chapter 3.5.1 --- Gene cassettes for carrot transformation --- p.58 / Chapter 3.5.1.1 --- Construction of gene cassettes for chicken or zebrafish bcmo driven by CaMV 35S promoter --- p.58 / Chapter 3.5.1.2 --- Construction of gene cassettes for chicken or zebrafish bcmo driven by lycopene-β-cyclase promoter --- p.63 / Chapter 3.5.2 --- Gene cassettes for Arabidopsis transformation --- p.67 / Chapter 3.5.2.1 --- Construction of gene cassettes expressing Dcpsy --- p.67 / Chapter 3.5.2.2 --- Construction of gene cassettes expressing mbcmos --- p.69 / Chapter 3.5.3 --- Gene cassettes for Rice transformation --- p.72 / Chapter 3.5.3.1 --- Construction of gene cassettes expressing mbcmos --- p.72 / Chapter 3.5.3.2 --- Construction of gene cassettes expressing Ospsyl and mbcmos --- p.74 / Chapter 3.5.4 --- Confirmation of sequence fidelity --- p.76 / Chapter 3.6 --- Carrot transformation --- p.76 / Chapter 3.6.1 --- Plant materials --- p.76 / Chapter 3.6.2 --- Preparation of Agrobacterium --- p.76 / Chapter 3.6.3 --- Agrobacterium mediated transformation --- p.77 / Chapter 3.6.3.1 --- Seed germination --- p.78 / Chapter 3.6.3.2 --- Co-cultivation with hypocotyls --- p.78 / Chapter 3.6.3.3 --- Callus induction and selection --- p.78 / Chapter 3.6.3.4 --- Liquid cell culture preparation and embryogenesis induction --- p.79 / Chapter 3.6.3.5 --- Regeneration --- p.80 / Chapter 3.7 --- Arabidopsis Transformation --- p.80 / Chapter 3.7.1 --- Plant materials --- p.80 / Chapter 3.7.2 --- Preparation of Agrobacterium --- p.81 / Chapter 3.7.3 --- Agrobacterium mediated transformation --- p.81 / Chapter 3.7.3.1 --- Co-cultivation --- p.81 / Chapter 3.7.3.2 --- Selection --- p.82 / Chapter 3.8 --- Rice transformation --- p.83 / Chapter 3.8.1 --- Plant materials --- p.83 / Chapter 3.8.2 --- Preparation of Agrobacterium --- p.83 / Chapter 3.8.3 --- Agrobacterium mediated transformation --- p.83 / Chapter 3.8.3.1 --- Callus induction from mature rice seeds --- p.84 / Chapter 3.8.3.2 --- Co-cultivation and selection --- p.84 / Chapter 3.9 --- Detection of transgene expression --- p.86 / Chapter 3.9.1 --- Detection at DNA level --- p.86 / Chapter 3.9.1.1 --- Genomic DNA extraction --- p.86 / Chapter 3.9.1.2 --- PCR screening --- p.86 / Chapter 3.9.1.3 --- Synthesis of DIG-labelled DNA probes --- p.86 / Chapter 3.9.1.4 --- Southern blot analysis --- p.87 / Chapter 3.9.2 --- Detection at RNA level --- p.88 / Chapter 3.9.2.1 --- Total RNA extraction --- p.88 / Chapter 3.9.2.2 --- Northern blot analysis --- p.89 / Chapter 3.9.2.3 --- RT-PCR --- p.89 / Chapter 3.9.3 --- Detection at protein level --- p.89 / Chapter 3.9.3.1 --- Antibody production --- p.89 / Chapter 3.9.3.1.1 --- B.CMO protein induction in pET30a-bacterial system --- p.90 / Chapter 3.9.3.1.2 --- Immunization of rabbit and serum collection --- p.93 / Chapter 3.9.3.2 --- Protein extraction and Tricine SDS-PAGE --- p.93 / Chapter 3.9.3.3 --- Western blot analysis --- p.94 / Chapter 3.9.4 --- Detection at final product level --- p.95 / Chapter 3.9.4.1 --- UPLC analysis --- p.95 / Chapter 3.9.4.1.1 --- Extraction of total carotenoids and retinoids --- p.95 / Chapter 3.9.4.1.2 --- UPLC identification --- p.96 / Chapter CHAPTER 4. --- RESULTS --- p.97 / Chapter 4.1 --- Modified bcmo genes --- p.97 / Chapter 4.2 --- Expression of BCMOs in bacterial system --- p.102 / Chapter 4.2.1 --- lac promoter system --- p.104 / Chapter 4.2.2 --- pBAD-TOPO® system --- p.106 / Chapter 4.2.3 --- UPLC detection --- p.108 / Chapter 4.3 --- Carrot transformation --- p.110 / Chapter 4.3.1 --- Construction of gene cassettes for carrot transformation --- p.110 / Chapter 4.3.2 --- Seed germination and co-cultivation --- p.112 / Chapter 4.3.3 --- Callus induction and selection --- p.113 / Chapter 4.3.4 --- Embryogenesis induction and regeneration --- p.113 / Chapter 4.3.5 --- Callus induction in the dark --- p.115 / Chapter 4.3.6 --- Detection of native BCMO --- p.116 / Chapter 4.3.6.1 --- Genomic PCR screening of 35Spro - zebcmo transgenic lines --- p.116 / Chapter 4.3.6.2 --- Southern blot analysis of 35Spro - zebcmo transgenic lines --- p.117 / Chapter 4.3.6.3 --- RT-PCR of 35Spro - zebcmo transgenic lines --- p.118 / Chapter 4.3.6.4 --- Detection at protein level --- p.119 / Chapter 4.3.6.4.1 --- Antibody production --- p.119 / Chapter 4.3.6.5 --- Western blot analysis of 35Spro - zebcmo transgenic lines --- p.123 / Chapter 4.3.6.6 --- Genomic PCR screening of later transgenic lines --- p.123 / Chapter 4.3.6.7 --- Western blot analysis of later transgenic lines --- p.125 / Chapter 4.3.6.8 --- UPLC analysis of later transgenic lines --- p.127 / Chapter 4.3.7 --- Detection of modified BCMO --- p.130 / Chapter 4.3.7.1 --- Genomic PCR screening --- p.130 / Chapter 4.3.7.2 --- Northern blot analysis --- p.132 / Chapter 4.3.7.3 --- Western blot analysis --- p.134 / Chapter 4.3.8 --- UPLC analysis --- p.136 / Chapter 4.4 --- Arabidopsis transformation --- p.138 / Chapter 4.4.1 --- Construction of gene cassettes for Arabidopsis transformation --- p.138 / Chapter 4.4.2 --- Selection --- p.139 / Chapter 4.4.3 --- Genmoic PCR screening of Arabidopsis transformants --- p.140 / Chapter 4.4.4 --- UPLC analysis for Arabidopsis transformants --- p.142 / Chapter 4.5 --- Rice transformation --- p.144 / Chapter 4.5.1 --- Construction of gene cassettes for rice transformation --- p.144 / Chapter 4.5.2 --- "Callus induction from mature rice seeds, co-cultivation and selection" --- p.146 / Chapter 4.5.3 --- Genomic PCR screening of Rice transformants --- p.147 / Chapter 4.5.4 --- UPLC analysis of rice transformants --- p.149 / Chapter CHAPTER 5. --- DISCUSSION --- p.151 / Chapter 5.1 --- Bacterial expression of BCMO --- p.151 / Chapter 5.2 --- Analysis of BCMO in plants --- p.153 / Chapter 5.2.1 --- Carrot --- p.154 / Chapter 5.2.1.1 --- Expression of BCMO in carrot transformants --- p.154 / Chapter 5.2.1.2 --- UPLC analysis of carrot transformants --- p.155 / Chapter 5.2.2 --- Arabidopsis --- p.156 / Chapter 5.2.3 --- Rice --- p.158 / Chapter 5.3 --- Proposed explanation for the failure of retinal production --- p.159 / Chapter 5.3.1 --- Retinal sequestration --- p.160 / Chapter 5.3.2 --- Localization of BCMO --- p.161 / Chapter 5.4 --- Future prospects --- p.163 / Chapter CHAPTER 6. --- CONCLUSIONS --- p.165 / REFERENCES --- p.166 / APPENDICES --- p.176

As economies develop, novel products are created and markets for these products arise. Genetically modified food (GMF) is an example of such a novel product and provides economists with the opportunity to investigate an infant market. Of particular interest with GMF is the impact of consumer reactions on the market. The response of consumers to GMF and their willingness to pay for it has emerged as an important factor in the development of this technology. This research investigates these consumer responses. Prior research suggests that two aspects of consumer behaviour may be relevant for the GMF market. First, consumers may react differently to different types of GMF, so that some products are potentially more economically viable. Secondly, some consumers appear to prefer not having GMF at all. Consumer behaviour is often framed according to neoclassical economic theory. Consumer preferences over goods and the attributes of those goods are generally held to have certain properties. The aspects of consumers' reactions to GMF noted above, however, may be in conflict with two properties of preferences in neoclassical theory. First, preferences over food attributes are not separable, but may interact with each other. Secondly, some consumers may have preferences regarding GMF that are not continuous. As a result, aggregate impacts of introducing GMF may be difficult to measure, which raises a third issue for investigation, aggregation. Finally, an alternative model of consumer behaviour is bounded rationality, which theorises that choices may be discontinuous as a result of specific protocols. It also suggests that consumers seek to make good-enough choices, rather than attempting to maximise their satisfaction. Thus, optimisation or maximisation is the fourth issue considered in this thesis. In order to investigate these properties of consumers' preferences, a choice experiment survey was developed. The strength of a choice experiment for examining these issues is its focus on the impact of each product attribute on a respondent's choices. Thus, it may be possible to identify potentially discontinuous choice patterns and to identify choices affected by interactions between GM technology and other food attributes. Results from a neoclassical analysis of the survey data suggest that some consumers consider the type of benefit created with GM technology in making their choices. In addition, one-quarter to one-half of respondents may have had discontinuous preferences with respect to GMF. Reactions to GMF appear related to respondents' attitudes, but not to socio-economic or demographic descriptors. As a result, aggregate measures of the impact of GMF may not fully account for consumers' responses. A boundedly rational model also has reasonable goodness of fit, and may provide a different perspective on consumer behaviour. It is hoped that the results of this research provide a better understanding of consumer behaviour regarding GMF and, by extension, of the process of consumer adoption of novel products. It is further hoped that this attempt to incorporate choice protocols into discrete choice analysis will provide a useful example for further research.

No description available.

<p>This study examines the possible incompatibility between environmental conventions and agreements promoting international trade. More specifically, it aims at discussing the Precautionary Principle and the WTO agreements on trade and how they might contradict each other on the international arena. The study focuses on the international trade with genetically modified organisms and discusses probable needs of reformulating the principles and/or conventions concerned if these are to function as tools of equal importance when handling environmental problems through policy making. The controversy is illustrated through a case-study of a trade dispute where the conflicting principles: the Precautionary Principle and WTO agreements on trade are involved, namely a dispute where a formal complaint to the WTO by the US, Canada and Argentina about the European Community supposedly applying a de facto moratorium on genetically modified organisms had the final outcome of the European Community being found to contravene international trade rules. The study concludes that actors on the international arena are still opinions apart concerning the way to perceive possible risks connected to biotechnology and genetically modified organisms. This incongruence is also found to apply to the status of the Precautionary Principle as a rule of international law as well as to which of the conventions that were applicable to the concerned dispute.</p>

<p>The aim was to study consumer attitudes to and perceptions of organic and genetically modified (GM) foods and factors influencing the purchase/non purchase of these two types of foods. Data were collected by two questionnaire surveys (random nation-wide samples of 2000 consumers, response rate 58% and 39% respectively) and one interview study (n=40).</p><p>A majority of the consumers had positive attitudes towards purchasing organic foods but few reported purchasing organic foods regularly. The most common beliefs about organic foods were that they are more expensive and healthier. Choice of organic foods was related to the perceived consequences for human health and to environmentally friendly behaviour. Perceived positive health consequences appeared to be a stronger motive for purchasing organic foods than were environmental benefits.</p><p>Attitudes towards genetic engineering (GE) appear to be dependent on the application area. Medical applications were generally more easily accepted than were food and agricultural applications. However, all medical applications were not equally accepted. Consumers had generally rather negative attitudes towards GM foods, which were perceived as not being healthy. GM of animals for food production was perceived more negative than GM of plants. Tangible benefits of GM foods, like being better for the environment or healthier, seemed to have a potential to increase willingness to buy.</p><p>The results indicate that the attitudes to organic and GM foods differ: the attitude to organic foods is positive while the attitude to GM foods is more negative. Further, the perceptions of organic and GM foods are opposed in relation to health. The purchase of organic foods and the potential willingness to purchase GM foods appear to be related to positive consequences for health and the environment.</p>

<p>The purpose of this thesis is to analyse the Swedish debate regarding genetically modified food products. What does the moral disagreements within this debate entail? In which way would it be possible to obtain and maintain a true dialogue? Is consensus a plausible or even a desirable goal? Can communicative ethics contribute? These questions are analysed in this research project.</p><p>The project focuses on three themes of the debate on genetically modified food 1990-2000 in Sweden. The first theme is in relation to the concepts: natural and unnatural. Within the debate on genetically modified food, there have been various wide ranging arguments on whether or not genetic modification is natural or unnatural. </p><p>The second theme concerns the concepts of risk and benefit. Safety questions and possible risks and benefits have been subjects of discussion and debate for a long time. This analysis highlights the way risks or benefits are valued. What does it mean if something is a risk? </p><p>The third theme concerns the argumentations regarding democracy, mostlyin relation to labelling discussions. Democracy is used to describe freedom of choice, another argumentation focuses on the right to be a part of the decision making process.</p><p>The last part of the thesis discusses the question whether communicative ethics can contribute to obtain dialogue between various parties, and whether consensus is possible or even a desirable as a goal. How would this function in a debate where there is an ethical disagreement based on divergent ethical standpoints? </p>

The purpose of this thesis is to analyse the Swedish debate regarding genetically modified food products. What does the moral disagreements within this debate entail? In which way would it be possible to obtain and maintain a true dialogue? Is consensus a plausible or even a desirable goal? Can communicative ethics contribute? These questions are analysed in this research project. The project focuses on three themes of the debate on genetically modified food 1990-2000 in Sweden. The first theme is in relation to the concepts: natural and unnatural. Within the debate on genetically modified food, there have been various wide ranging arguments on whether or not genetic modification is natural or unnatural. The second theme concerns the concepts of risk and benefit. Safety questions and possible risks and benefits have been subjects of discussion and debate for a long time. This analysis highlights the way risks or benefits are valued. What does it mean if something is a risk? The third theme concerns the argumentations regarding democracy, mostlyin relation to labelling discussions. Democracy is used to describe freedom of choice, another argumentation focuses on the right to be a part of the decision making process. The last part of the thesis discusses the question whether communicative ethics can contribute to obtain dialogue between various parties, and whether consensus is possible or even a desirable as a goal. How would this function in a debate where there is an ethical disagreement based on divergent ethical standpoints?

The aim was to study consumer attitudes to and perceptions of organic and genetically modified (GM) foods and factors influencing the purchase/non purchase of these two types of foods. Data were collected by two questionnaire surveys (random nation-wide samples of 2000 consumers, response rate 58% and 39% respectively) and one interview study (n=40). A majority of the consumers had positive attitudes towards purchasing organic foods but few reported purchasing organic foods regularly. The most common beliefs about organic foods were that they are more expensive and healthier. Choice of organic foods was related to the perceived consequences for human health and to environmentally friendly behaviour. Perceived positive health consequences appeared to be a stronger motive for purchasing organic foods than were environmental benefits. Attitudes towards genetic engineering (GE) appear to be dependent on the application area. Medical applications were generally more easily accepted than were food and agricultural applications. However, all medical applications were not equally accepted. Consumers had generally rather negative attitudes towards GM foods, which were perceived as not being healthy. GM of animals for food production was perceived more negative than GM of plants. Tangible benefits of GM foods, like being better for the environment or healthier, seemed to have a potential to increase willingness to buy. The results indicate that the attitudes to organic and GM foods differ: the attitude to organic foods is positive while the attitude to GM foods is more negative. Further, the perceptions of organic and GM foods are opposed in relation to health. The purchase of organic foods and the potential willingness to purchase GM foods appear to be related to positive consequences for health and the environment.

This study examines the possible incompatibility between environmental conventions and agreements promoting international trade. More specifically, it aims at discussing the Precautionary Principle and the WTO agreements on trade and how they might contradict each other on the international arena. The study focuses on the international trade with genetically modified organisms and discusses probable needs of reformulating the principles and/or conventions concerned if these are to function as tools of equal importance when handling environmental problems through policy making. The controversy is illustrated through a case-study of a trade dispute where the conflicting principles: the Precautionary Principle and WTO agreements on trade are involved, namely a dispute where a formal complaint to the WTO by the US, Canada and Argentina about the European Community supposedly applying a de facto moratorium on genetically modified organisms had the final outcome of the European Community being found to contravene international trade rules. The study concludes that actors on the international arena are still opinions apart concerning the way to perceive possible risks connected to biotechnology and genetically modified organisms. This incongruence is also found to apply to the status of the Precautionary Principle as a rule of international law as well as to which of the conventions that were applicable to the concerned dispute.

A great effort has been recently devoted to the development of new devices for the detection of specific sequences of DNA, due to increasing need of label - free, fast, cheap, and miniaturized analytical systems able to detect target sequences for screening purposes, especially in food industry for genetically modified organisms (GMOs). In this study, development of a QCM - based DNA biosensor for the detection of the hybridisation of CaMV 35S promoter sequence (P35S) was investigated. Attention was focused on the choice of the coating chemistry that could be used for the immobilisation of probe sequences on the gold surface of the quartz crystal. Two immobilisation procedures were tested and compared considering the amount of the immobilised probe, the extent of the hybridisation reaction, the possibility of regeneration and the absence of non - specific adsorption. The two coating methods were based on the use of self - assembled monolayers. One of them employed the interaction between the thiol and gold for the immobilisation of a thiolated P35S probe, while the other employed formation of functionalised aldehyde groups by ethylenediamine plasma polymerization on the gold surface for the immobilisation of amined P35S probes through gluteraldehyde activation. Results indicated that immobilisation of a thiolated probe provides better immobilisation characteristic, higher sensitivity for the detection of the hybridisation reaction, absence of non - specific adsorption and a higher stability with respect to the regeneration step. The optimised immobilisation procedure for the thiolated probe was used for the detection of P35S sequence in PCR - amplified DNAs and in real samples of pflp - gene inserted tobacco plants that produce ferrodoxin like protein additionally. Fragmentation of the genomic DNAs were achieved by digestion with restriction endonucleases and sonication. The obtained results from the fragmented genomic DNAs demonstrated that it is possible to detect the target sequence directly in non-amplified genomic DNAs by using the developed QCM - based DNA biosensor system. The developed QCM-based DNA biosensor represented promising results for a real-time, label - free, direct detection of DNA samples for the screening of GMOs.