Fung Ka Lee. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2003. / Includes bibliographical references (leaves 159-179). / Abstracts in English and Chinese. / ACKNOWLEDGEMENTS --- p.II / ABBREVIATIONS --- p.III / ABSTRACTS --- p.V / LIST OF FIGURES --- p.VIII / LIST OF TABLES --- p.IX / CONTENTS --- p.X / CONTENTS / Chapter CHAPTER ONE: --- GENERAL INTRODUCTION / Chapter 1.1. --- Review: Cellular mechanisms of drug resistance in cancer cells --- p.2 / Chapter 1.1.1. --- Drug resistance by decrease of drug accumulation --- p.2 / Chapter 1.1.2. --- Drug resistance conferred by detoxification of drug in cells --- p.5 / Chapter 1.1.3. --- Drug resistance conferred by alteration of drug targets or by enhancement of target repair --- p.6 / Chapter 1.1.4. --- Drug resistance conferred by genes alteration in apoptotic pathways --- p.9 / Chapter 1.2. --- Objective and review --- p.11 / Chapter CHAPTER TWO: --- PROFILING AND IDENTIFICATION OF DIFFERENTIALLY EXPRESSED GENES OF HUMAN SQUAMOUS CARCINOMA A431 CELLS BY mRNA DIFFERENTIAL DISPLAY AND cDNA MICROARRAY / Chapter 2.1. --- Introduction / Chapter 2.1.1. --- Changes in gene expression and drug resistance --- p.16 / Chapter 2.1.2. --- Current states of technologies in gene expression analysis --- p.17 / Chapter 2.1.3. --- Monitoring the gene expression profile in the field of drug resistance by cDNA microarray --- p.18 / Chapter 2.1.4. --- Monitoring the gene expression profile in the field of drug resistance by differential display --- p.22 / Chapter 2.2. --- Materials and Methods / Chapter 2.2.1. --- Materials --- p.27 / Chapter 2.2.2. --- Methods / Chapter 2.2.2.1. --- Cell culture and cell lines --- p.29 / Chapter 2.2.2.2. --- cDNA microarray / Chapter 2.2.2.2.1. --- RNA preparation --- p.29 / Chapter 2.2.2.2.2. --- Differential hybridization of ResGen GenFilters Mammalian micro array --- p.30 / Chapter 2.2.2.3. --- mRNA differential display / Chapter 2.2.2.3.1. --- RNA preparation --- p.31 / Chapter 2.2.2.3.2. --- RT-PCR based mRNA differential display --- p.31 / Chapter 2.2.2.3.3. --- Reamplification of cDNA probes --- p.32 / Chapter 2.2.2.3.4. --- Subcloning of differentially expressed cDNAs / Chapter A. --- Preparation of ultra-competent E.coli cells for transformation --- p.33 / Chapter B. --- Preparation of cloning vector and DNA transformation --- p.33 / Chapter 2.2.2.3.5. --- Verification of cDNA expression by colony-PCR and cDNA probes / Chapter A. --- colony-PCR --- p.34 / Chapter B. --- Bacterial glycerol stock and plasmid preparation --- p.34 / Chapter C. --- cDNA probes preparation for Northern blot analysis --- p.35 / Chapter 2.2.2.3.6. --- Northern blot analysis --- p.35 / Chapter 2.2.2.3.7. --- Sequencing of cDNA cloned inserts --- p.36 / Chapter 2.3. --- Results / Chapter 2.3.1. --- Differential gene expression profile of A431 cells --- p.43 / Chapter 2.3.2. --- Identification of differentially expressed genes by mRNA differential display --- p.53 / Chapter 2.4. --- Discussion --- p.60 / Chapter CHAPTER THREE: --- CHARACTERIZATION OF DIFFERENTIALLY EXPRESSED GENES.。PART 1: THE ROLE OF VACUOLAR PROTON PUMP IN REGULATION OF DRUG SENSITIVITY AND APOPTOSIS IN A431 CELLS / Chapter 3.1. --- Introduction / Chapter 3.1.1. --- An overview of vacuolar proton pump/vacuolar-H+-ATPase (V-ATPase): Structure and function --- p.66 / Chapter 3.1.2. --- V-ATPases and drug resistance mechanisms --- p.69 / Chapter 3.1.3. --- Objective --- p.70 / Chapter 3.2. --- Materials and Methods / Chapter 3.2.1. --- Materials --- p.73 / Chapter 3.2.2. --- Methods / Chapter 3.2.2.1. --- Cell culture and cell lines --- p.74 / Chapter 3.2.2.2. --- "RNA isolation, mRNA differential display, cDNA reamplification and colony-PCR verification" --- p.74 / Chapter 3.2.2.3. --- Cellular pH measurement by flow cytometry --- p.75 / Chapter 3.2.2.4. --- MTT Drug Sensitivity Assay --- p.76 / Chapter 3.2.2.5. --- DNA fragmentation assay --- p.77 / Chapter 3.3. --- Results / Chapter 3.3.1. --- Identification of cDNA band encoding vacuolar proton pump subunit c by differential display --- p.78 / Chapter 3.3.2. --- Effect of bafilomycin A1 (BAF) and concanamycin A (CON) on DOX- and CP-induced cytotoxicity and apoptosis ' --- p.79 / Chapter 3.3.3. --- Elevation of cellular pH in cisplatin-resistant cells correlated with cisplatin resistance mechanism of A431 cells --- p.94 / Chapter 3.4. --- Discussion --- p.98 / Chapter CHAPTER FOUR: --- CHARACTERIZATION OF DIFFERENTIALLY EXPRESSED GENES。PART 2: IDENTIFICATION OF A NOVEL cDNA OVEREXPRESSED IN HUMAN SQUAMOUS CARCINOMA A431 PARENT CELLS / Chapter 4.1. --- Introduction --- p.102 / Chapter 4.2. --- Materials and Methods / Chapter 4.2.1. --- Materials --- p.105 / Chapter 4.2.2. --- Methods / Chapter 4.2.2.1. --- Cell culture and cell lines --- p.105 / Chapter 4.2.2.2. --- "RNA isolation, mRNA differential display, cDNA reamplification and colony-PCR verification" --- p.105 / Chapter 4.2.2.3. --- Northern blot analysis --- p.106 / Chapter 4.2.2.4. --- Human tissue distribution of clone PA-8P19 --- p.107 / Chapter 4.3. --- Results / Chapter 4.3.1. --- Identification of novel cDNA clone PA-8P19 in A431 cells by differential display --- p.108 / Chapter 4.3.2. --- Gene expression profile of clone PA-8P19 in human tissues and other tumor cell lines --- p.113 / Chapter 4.3.3. --- Regulation of PA-8P19 expression by DNA damaging agents --- p.117 / Chapter 4.4. --- Discussion --- p.122 / Chapter CHAPTER FIVE: --- PROFILING OF DIFFERENTIAL EXPRESSED PROTEINS BY TWO-DIMENSIONAL GEL ELECTPOPHORESIS / Chapter 5.1. --- Introduction / Chapter 5.1.1. --- Protein analysis on proteomic scale --- p.125 / Chapter 5.1.2. --- Protein expression profiling for studying drug resistance by Two- dimensional gel electrophoresis --- p.127 / Chapter 5.2. --- Materials and Methods / Chapter 5.2.1. --- Materials --- p.130 / Chapter 5.2.2. --- Methods / Chapter 5.2.2.1. --- Cell culture and cell lines --- p.132 / Chapter 5.2.2.2. --- Two-dimensional gel electrophoresis / Chapter I. --- Sample preparation --- p.132 / Chapter II. --- First-dimensional gel: isoelectric focusing (IEF) --- p.133 / Chapter III. --- Second-dimensional gel: Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) --- p.134 / Chapter IV. --- Protein visualization and image analysis --- p.134 / Chapter V. --- In-gel digestion --- p.135 / Chapter VI. --- Matrix-assisted laser desorption ionization Time-of-Flight Mass Spectrometry (MALDI-TOF-MS) and database search --- p.136 / Chapter 5.2.2.3. --- Western blot analysis --- p.137 / Chapter 5.3. --- Results / Chapter 5.3.1. --- Identification of differentially expressed proteins in A431 cells by two-dimensional gel electrophoresis --- p.140 / Chapter 5.4. --- Discussion / Chapter 5.4.1. --- Two-dimensional gel electrophoresis as a powerful tool for identification of differential protein expression in A431 cells --- p.150 / Chapter CHAPTER SIX: --- GENERAL CONCLUSION AND PERSPECTIVES --- p.155 / REFERENCES --- p.159
| Identifer | oai:union.ndltd.org:cuhk.edu.hk/oai:cuhk-dr:cuhk_324455 |
| Date | January 2003 |
| Contributors | Fung, Ka Lee., Chinese University of Hong Kong Graduate School. Division of Biochemistry. |
| Source Sets | The Chinese University of Hong Kong |
| Language | English, Chinese |
| Detected Language | English |
| Type | Text, bibliography |
| Format | print, xiii, 179 leaves : ill. (some col.) ; 30 cm. |
| Rights | Use of this resource is governed by the terms and conditions of the Creative Commons “Attribution-NonCommercial-NoDerivatives 4.0 International” License (http://creativecommons.org/licenses/by-nc-nd/4.0/) |
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