Thesis (Ph. D.)--University of Wisconsin--Madison, 1982. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references.

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Tübingen, Universiẗat, Diss., 2004.

Thesis (Ph. D.)--West Virginia University, 2003. / Title from document title page. Document formatted into pages; contains vi, 110 p. : ill. Includes abstract. Includes bibliographical references.

Thesis (Ph. D.)--University of Hong Kong, 2008. / Also available in print.

Chan Chung Yiu Patrick. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2004. / Includes bibliographical references (leaves 134-151). / Abstracts in English and Chinese. / Abstract --- p.i / 摘要 --- p.iii / Acknowledgements --- p.v / Table of Contents --- p.vi / List of Tables --- p.ix / List of Figures --- p.x / Abbreviations --- p.xv / Chapter CHAPTER 1 --- Literature review --- p.1 / Chapter 1.1 --- Aquatic heavy metal contaminations --- p.1 / Chapter 1.1.1 --- Biology of heavy metals --- p.1 / Chapter 1.1.2 --- Mechanism of heavy metal toxicity --- p.2 / Chapter 1.2 --- Environmental monitoring of aquatic heavy metal contaminations --- p.4 / Chapter 1.2.1 --- Bioconcentration effects of heavy metals --- p.4 / Chapter 1.2.2. --- The concept of bioindicator in pollution assessment --- p.5 / Chapter 1.3 --- Metallothioneins --- p.7 / Chapter 1.3.1 --- Biological functions of MT and its metal inducibility --- p.7 / Chapter 1.4 --- Zebrafish (Daino reio) as an animal model --- p.11 / Chapter 1.4.1 --- Biology of zebrafish --- p.11 / Chapter 1.4.2 --- Current applications of transgenic zebrafish --- p.12 / Chapter 1.5 --- The use of cell culture systems in toxicology research --- p.13 / Chapter 1.6 --- Project aims --- p.16 / Chapter CHAPTER 2 --- Quantification of zMT mRNA levels using real-time PCR --- p.17 / Chapter 2.1 --- Introduction --- p.17 / Chapter 2.1.1 --- The use of zebrafish embryos in toxicity assessment --- p.17 / Chapter 2.1.2 --- MT mRNA as a bioindicator of metal exposure --- p.18 / Chapter 2.1.3 --- Quantification of gene transcripts by RT-PCR --- p.19 / Chapter 2.1.4 --- Specific aims of this chapter --- p.27 / Chapter 2.2 --- Materials and methods --- p.28 / Chapter 2.2.1 --- Animal --- p.28 / Chapter 2.2.2 --- Cell culture --- p.31 / Chapter 2.2.3 --- General molecular biology techniques --- p.33 / Chapter 2.2.4 --- mRNA quantification by Real-time PCR --- p.35 / Chapter 2.3 --- Results --- p.42 / Chapter 2.3.1 --- Heavy metal toxicity --- p.42 / Chapter 2.3.1.1 --- In vivo metal toxicity in zebrafish adult --- p.43 / Chapter 2.3.1.2 --- In vivo metal toxicity in zebrafish embryos at late epiboly --- p.44 / Chapter 2.3.1.3 --- In vivo metal toxicity in zebrafish eleutheroembryo --- p.46 / Chapter 2.3.1.4 --- In vitro metal toxicity on ZFL cell line --- p.48 / Chapter 2.3.2 --- Optimization of real-time PCR conditions --- p.50 / Chapter 2.3.2.1 --- Construction of relative standard curve --- p.50 / Chapter 2.3.2.2 --- Optimization of primer concentration --- p.52 / Chapter 2.3.2.3 --- Melting curve analysis for PCR specificity --- p.53 / Chapter 2.2.3 --- Quantification of zMT mRNA by Real-time PCR --- p.54 / Chapter 2.2.3.1 --- Relative zMT mRNA induction in zebrafish embryos at late epiboly --- p.54 / Chapter 2.2.3.2 --- Relative zMT mRNA induction in zebrafish eleutherombryos --- p.57 / Chapter 2.2.3.3 --- Relative zMT mRNA induction in SJD. 1 cell line --- p.58 / Chapter 2.2.2.4 --- In vitro zMT mRNA induction in ZFL cell line --- p.62 / Chapter 2.4 --- Discussions --- p.64 / Chapter 2.4.1 --- Change in metal sensitivity during zebrafish embryo development --- p.64 / Chapter 2.4.2 --- Developmental stage-specfic inducibility of zMT gene expression and metal toxicity --- p.65 / Chapter 2.4.3 --- In vitro zMT regulation by heavy metal ions --- p.67 / Chapter 2.4.4 --- The potential use of zMT expression as exposure biomarker --- p.69 / Chapter CHAPTER 3 --- Functional analysis of a cloned zMT-II gene promoter in zebrafish cell-lines: SJD.1 and ZFL --- p.71 / Chapter 3.1 --- Zebrafish MT gene promoter --- p.71 / Chapter 3.1.1 --- The structure of zMT promoter --- p.71 / Chapter 3.1.2 --- Functional analysis of cloned zMT-II promoter region in HepG2 cells --- p.72 / Chapter 3.1.3 --- Specific aims of this chapter --- p.77 / Chapter 3.2 --- Materials and methods --- p.78 / Chapter 3.2.1 --- General molecular biology techniques --- p.78 / Chapter 3.2.2 --- Cell culture --- p.79 / Chapter 3.2.3 --- Transient transfection assay --- p.81 / Chapter 3.3 --- Results --- p.84 / Chapter 3.3.1 --- Metal responsiveness of zMT-II promoter by transient transfection --- p.84 / Chapter 3.3.2 --- Deletion analysis --- p.88 / Chapter 3.3.2.1 --- Deletion analysis of zMT-II gene promoter in SJD. 1 cell line --- p.88 / Chapter 3.3.2.2 --- Deletion analysis of zMT-II gene promoter in ZFL cell line --- p.90 / Chapter 3.4 --- Discussions --- p.91 / Chapter 3.4.1 --- Structure of zMT-II gene promoter --- p.91 / Chapter 3.4.2 --- Metal responsiveness of zMT-II promoter --- p.94 / Chapter 3.4.3 --- Deletion analysis of zMT-IIgene promoter --- p.95 / Chapter CHAPTER 4 --- Transgenic zebrafish model for in vivo zMT gene regulation study --- p.97 / Chapter 4.1 --- Introduction --- p.97 / Chapter 4.1.1 --- Development of transgenic fish --- p.97 / Chapter 4.1.2 --- The principle of gene delivery --- p.98 / Chapter 4.1.3 --- The application of transgenic zebrafish model --- p.99 / Chapter 4.1.4 --- Specific aim of this chapter --- p.101 / Chapter 4.2 --- Materials and methods --- p.102 / Chapter 4.2.1 --- General molecular biology techniques --- p.102 / Chapter 4.2.2 --- Sequence of primers used --- p.103 / Chapter 4.2.3 --- Engineering of constructs for transgenic zebrafish study --- p.103 / Chapter 4.2.4 --- In vitro efficacy test of the GFP constructs --- p.105 / Chapter 4.2.5 --- Gene transfer into zebrafish embryos by electroporation --- p.107 / Chapter 4.2.6 --- Screening of transgenic candidates --- p.108 / Chapter 4.3 --- Results --- p.110 / Chapter 4.3.1 --- Engineering of DNA constructs for transgenic zebrafish production --- p.110 / Chapter 4.3.2 --- In vitro efficacy test of the DNA constructs --- p.111 / Chapter 4.3.3 --- Optimization of electroporation voltage --- p.117 / Chapter 4.4.4 --- Screening of transgenic candidates --- p.118 / Chapter 4.4 --- Discussion --- p.120 / Chapter 4.4.1 --- Potential application of the zMT promoter transgenic mode --- p.120 / Chapter 4.4.2 --- The use of GFP transgenic zebrafish model in developmental gene regulation study --- p.121 / Chapter 4.4.3 --- In vitro efficacy of the GFP constructs --- p.122 / Chapter 4.4.4 --- Transgene expression in zebrafish --- p.122 / Chapter CHAPTER 5 --- General discussion --- p.126 / REFERENCES --- p.134

by Cheung Pok Lap. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2003. / Includes bibliographical references (leaves 225-251). / Abstracts in English and Chinese. / Abstract --- p.i / 摘錄 --- p.iii / Acknowledgements --- p.v / Table of Contents --- p.vi / List of Tables --- p.ix / List of Figures --- p.x / Abbreviations --- p.xiv / Chapter CHAPTER 1 --- LITERATURE REVIEW --- p.1 / Chapter 1.1 --- Biology of Metals --- p.1 / Chapter 1.1.1 --- Mechanism for Monitoring and Controlling Intracellular Metal Ions --- p.2 / Chapter 1.1.2 --- Metal Ions Homeostasis --- p.5 / Chapter 1.2 --- Metallothionein (MT) --- p.8 / Chapter 1.2.1 --- Classification of MT --- p.8 / Chapter 1.2.2 --- Structure of MT --- p.9 / Chapter 1.2.3 --- Organization of MT Genes --- p.11 / Chapter 1.2.4 --- Biological Functions of MT --- p.13 / Chapter 1.2.5 --- MT as a model of Transcriptional Regulation of Gene Expression --- p.19 / Chapter 1.3 --- Fish MT Genes --- p.24 / Chapter 1.4 --- Aims and Rationale of Present Study --- p.28 / Chapter CHAPTER 2 --- CLONING AND CHARACTERIZATION OF TILAPIA MT (tiMT) GENE PROMOTERS --- p.30 / Chapter 2.1 --- Introduction / Chapter 2.1.1 --- The Biology of Tilapia --- p.30 / Chapter 2.1.2 --- The Study of Tilapia MT --- p.31 / Chapter 2.1.3 --- Fish MT Promoters --- p.35 / Chapter 2.1.4 --- Specific Aims of This Chapter --- p.36 / Chapter 2.2 --- Materials and methods --- p.37 / Chapter 2.2.1 --- Animals --- p.37 / Chapter 2.2.2 --- General Molecular Biology Technique --- p.37 / Chapter 2.2.3 --- PCR Primers Used --- p.40 / Chapter 2.2.4 --- Cloning of tilapia MT Gene 5'-flanking Region Using Inverse PCR --- p.41 / Chapter 2.2.5 --- Cloning of full length of tilapia MT Genes / Chapter 2.2.6 --- Transient Transfection Assay --- p.43 / Chapter 2.3 --- Results --- p.47 / Chapter 2.3.1 --- Tilapia MT Genes --- p.47 / Chapter 2.3.2 --- Functional Analysis of tiMT Gene Promoter by Transient Transfection --- p.49 / Chapter 2.4 --- Discussions --- p.58 / Chapter 2.4.1 --- Tilapia MT Genes --- p.58 / Chapter 2.4.2 --- Functional Analysis of tiMT Gene Promoter by Transient Transfection --- p.63 / Chapter 2.5 --- Conclusion --- p.67 / Chapter CHAPTER 3 --- DETECTION OF MT mRNA LEVELS BY QUANTITATIVE RT-COMPETITIVE PCR --- p.68 / Chapter 3.1 --- Introduction --- p.68 / Chapter 3.1.1 --- Quantitative RT-competitive PCR --- p.69 / Chapter 3.1.2 --- Specific Aims of This Chapter --- p.71 / Chapter 3.2 --- Materials and methods --- p.72 / Chapter 3.2.1 --- Animals --- p.72 / Chapter 3.2.2 --- Isolation of total RNA and preparation of first strand cDNA --- p.72 / Chapter 3.2.3 --- Design of primers for preparation of MT mimic and competitive PCR --- p.73 / Chapter 3.2.4 --- Preparation of MT mimic cDNA --- p.74 / Chapter 3.2.5 --- Optimization PCR cycle number for β-actin and MT amplification --- p.77 / Chapter 3.2.6 --- Quantitative competitive PCR --- p.78 / Chapter 3.2.7 --- Quantitative analysis --- p.81 / Chapter 3.2.8 --- Statistical analysis --- p.82 / Chapter 3.3 --- Results --- p.83 / Chapter 3.3.1 --- Preparation of MT mimic cDNA --- p.83 / Chapter 3.3.2 --- Optimization PCR cycle number for MT and β-actin amplification --- p.84 / Chapter 3.3.3 --- Quantification of the MT cDNA levels by competitive PCR --- p.86 / Chapter 3.4 --- Discussions --- p.154 / Chapter 3.4.1 --- Comparison of MT Gene Expression in both in vivo tilapia liver andin vitro PLHC-1 fish cell line --- p.154 / Chapter 3.4.2 --- Absolute Quantification of mRNA using Real Time RT-PCR --- p.159 / Chapter 3.5 --- Conclusion --- p.160 / Chapter CHAPTER 4 --- TILAPIA MTF-1: PCR-CLONING AND GENE EXPRESSION STUDIES --- p.161 / Chapter 4.1 --- Introduction --- p.161 / Chapter 4.1.1 --- General Features of MTF-1 --- p.164 / Chapter 4.1.2 --- Activation MRE-binding of the MTF-1 --- p.166 / Chapter 4.1.3 --- Possible Models for Heavy-metal Regulated MT Genes Transcription --- p.169 / Chapter 4.1.4 --- Genes Under the Regulation of MTF-1 --- p.175 / Chapter 4.1.5 --- Specific Aims of This Chapter --- p.176 / Chapter 4.2 --- Materials and methods --- p.177 / Chapter 4.2.1 --- Cloning of a partial fragment of MTF-1 in tilapia --- p.177 / Chapter 4.2.2 --- Rapid Amplification of cDNA 5'ends --- p.180 / Chapter 4.2.3 --- Rapid Amplification of cDNA 3'ends --- p.184 / Chapter 4.2.4 --- Cloning of The Full-Length Tilapia MTF-1 cDNA Isoforms --- p.185 / Chapter 4.2.5 --- Northern blot analysis of MTF-1 transcripts in tilapia tissues --- p.186 / Chapter 4.2.6 --- Differential Expression of MTF-1 Isoforms in Tilapia Tissues --- p.187 / Chapter 4.2.7 --- Cotransfection Study on Tilapia MTF-1 cDNAs --- p.189 / Chapter 4.3 --- Results --- p.191 / Chapter 4.3.1 --- PCR Amplification of the Partial Sequence of tiMTF-1 --- p.191 / Chapter 4.3.2 --- Rapid Amplification of cDNA 5' and 3' ends of tiMTF-1 --- p.193 / Chapter 4.3.3 --- Cloning of The Full-Length Tilapia MTF-1 cDNA Isoforms --- p.196 / Chapter 4.3.4 --- Northern Blot Analysis of tilapia MTF-1 isoforms --- p.205 / Chapter 4.3.5 --- Differential Expression of MTF-1 Isoforms in Tilapia Tissues --- p.206 / Chapter 4.3.6 --- Cotransfection Study on Tilapia MTF-1 cDNAs --- p.207 / Chapter 4.4 --- Discussions --- p.212 / Chapter 4.4.1 --- Tilapia MTF-1 --- p.212 / Chapter 4.4.2 --- Biological Activity of MTF-1 --- p.216 / Chapter 4.5 --- Conclusion --- p.218 / Chapter CHAPTER 5 --- GENERAL DISCUSSION --- p.220 / REFERENCES --- p.225

Chan, Wai Lun. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2007. / Includes bibliographical references (leaves 140-157). / Abstracts in English and Chinese. / Abstract --- p.i / 摘要 --- p.iii / Acknowledgements --- p.v / Table of Contents --- p.1 / List of Tables --- p.4 / List of Figures --- p.5 / List of Abbreviations --- p.8 / Chapter 1. --- Introduction --- p.10 / Chapter 1.1 --- Biology of metals --- p.10 / Chapter 1.2 --- Metal detoxification systems --- p.11 / Chapter 1.3 --- Metallothionein --- p.13 / Chapter 1.4 --- Classification of MTs --- p.15 / Chapter 1.5 --- Biological roles of MT --- p.15 / Chapter 1.5.1 --- Homeostasis of essential transition metal ion --- p.15 / Chapter 1.5.2 --- Detoxification of non-essential heavy metal ion --- p.17 / Chapter 1.5.3 --- Protection against oxidative stress --- p.18 / Chapter 1.5.4 --- Role in neurodegenerative diseases --- p.19 / Chapter 1.6 --- Molecular biology of MT --- p.19 / Chapter 1.6.1 --- MT gene structure --- p.19 / Chapter 1.6.2 --- MT gene regulation --- p.21 / Chapter 1.7 --- MRE binding transcription factor-1 (MTF-1) --- p.30 / Chapter 1.8 --- Activation of MTF-1 --- p.31 / Chapter 1.9 --- Target genes of MTF-1 --- p.32 / Chapter 1.10 --- Fish MT gene and MTF-1 --- p.33 / Chapter 1.11 --- Tilapia --- p.39 / Chapter 1.12 --- Study of tilapia MT --- p.41 / Chapter 1.13 --- Aims and rationale of study --- p.43 / Chapter 2. --- Materials and Methods --- p.45 / Chapter 2.1 --- Cloning of tilapia MT gene 5'-flanking region --- p.45 / Chapter 2.1.1 --- Animals --- p.45 / Chapter 2.1.2 --- Preparation of tilapia genomic DNA --- p.45 / Chapter 2.1.3 --- DNA walking --- p.45 / Chapter 2.1.4 --- Amplification of whole tiMT gene --- p.50 / Chapter 2.2 --- Determination of transcription start site --- p.51 / Chapter 2.2.1 --- Total RNA extraction --- p.51 / Chapter 2.2.2 --- Rapid amplification of 5，complementary DNA ends (5' RACE) --- p.52 / Chapter 2.3 --- Transient transfection assay --- p.54 / Chapter 2.3.1 --- Cell culture --- p.54 / Chapter 2.3.2 --- Construction of pGL3-tiMT deletion mutants --- p.54 / Chapter 2.3.3 --- Preparation of heavy metal solutions --- p.56 / Chapter 2.3.4 --- Determination of heavy metal ion toxicities by alamarBlue´ёØ assay --- p.56 / Chapter 2.3.5 --- Transient transfection of plasmids to Hepa-T1 cells --- p.56 / Chapter 2.3.6 --- Metal ions treatment and study of tiMT promoter activities --- p.57 / Chapter 2.3.7 --- Transient gene expression studies of deletion mutants of tiMT promoter --- p.57 / Chapter 2.4 --- Site-directed mutagenesis of tiMT promoter --- p.58 / Chapter 2.4.1 --- Polymerase chain reaction (PCR)-based site-directed mutagenesis --- p.58 / Chapter 2.4.2 --- Transient transfection of plasmids to Hepa-T1 cells and study of tiMT promoter activities --- p.62 / Chapter 2.5 --- Electrophoretic mobility shift assay (EMSA) --- p.63 / Chapter 2.5.1 --- Extract preparation --- p.63 / Chapter 2.5.2 --- Preparation of radiolabeled tiMRE oligonucleotides --- p.63 / Chapter 2.5.3 --- Electrophoretic mobility shift assay (EMSA) --- p.64 / Chapter 3. --- Results --- p.66 / Chapter 3.1 --- "Cloning of tilapia MT (tiMT) gene 5,-flanking region and amplification of whole tiMT gene" --- p.66 / Chapter 3.2 --- Determination of transcription start site --- p.69 / Chapter 3.3 --- Cloning of tiMT promoter fragment into reporter vector --- p.72 / Chapter 3.4 --- Determination of heavy metal ion toxicities by alamarBlue´ёØ assay --- p.72 / Chapter 3.5 --- Study of tiMT promoter activities by heavy metal ions exposure..… --- p.72 / Chapter 3.6 --- Cloning of deletion mutants of tiMT promoter --- p.79 / Chapter 3.7 --- Transient gene expression studies of deletion mutants of tiMT promoter --- p.80 / Chapter 3.8 --- Cloning of mutants with site-directed mutagenesis in tiMT promoter --- p.88 / Chapter 3.9 --- Site-directed mutagenesis of tiMT promoter --- p.92 / Chapter 3.10 --- Electrophoretic Mobility Shift Assay (EMSA) --- p.97 / Chapter 4. --- Discussion --- p.102 / Chapter 4.1 --- Tilapia MT gene --- p.102 / Chapter 4.2 --- Resistance of tilapia to heavy metal ions --- p.107 / Chapter 4.3 --- Functional analysis of tiMT gene promoter by transient transfection --- p.111 / Chapter 4.4 --- DNA binding of metal responsive transcription factor in Hepa-T1 cells --- p.121 / Chapter 4.5 --- Conclusion --- p.138 / References --- p.140

published_or_final_version / abstract / Chemistry / Doctoral / Doctor of Philosophy

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