Au Chi Chuen. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2002. / Includes bibliographical references (leaves 157-169). / Abstracts in English and Chinese. / Acknowledgement --- p.i / Table of Contents --- p.ii / Abstract --- p.v / 論文摘要 --- p.vii / Abbreviations --- p.viii / List of Figures --- p.ix / List of Tables --- p.x / Chapter Chapter 1 --- Introduction / Chapter 1.1 --- General introduction --- p.1 / Chapter 1.2 --- Hepatitis B virus and Hepatocellular carcinoma --- p.3 / Chapter 1.3 --- Pathogenesis of HBV related HCC --- p.6 / Chapter 1.4 --- Current screening test and tumor markers --- p.10 / Chapter 1.5 --- Expressed sequence tag (EST) sequencing --- p.13 / Chapter 1.6 --- Aim of the present study --- p.15 / Chapter 1.7 --- Characterization of KIAA0022 --- p.16 / Chapter Chapter 2 --- Materials and Methods / Chapter 2.1 --- Construction of liver HCC and normal counterpart libraries --- p.19 / Chapter 2.2 --- Plating out the human liver cDNA libraries --- p.19 / Chapter 2.3 --- PCR amplification of clones human liver cancer and the normal counterpart cDNA libraries --- p.21 / Chapter 2.4 --- Cycle sequencing of cloned human liver cancer and the normal counterpart cDNA libraries --- p.21 / Chapter 2.4.1 --- Dye-primer cycle sequencing (Pharmacia) --- p.21 / Chapter 2.4.1.1 --- Using Pharmacia LBKA.L.F. DNA sequencer --- p.21 / Chapter 2.4.1.2 --- Using Li-Cor 4200 Automated DNA sequencer --- p.22 / Chapter 2.4.2 --- Dye-terminator cycle sequencing (Pharmacia) --- p.22 / Chapter 2.5 --- Sequences analysis --- p.23 / Chapter 2.6 --- Cloning of full-length cDNA of KIAA0022 --- p.24 / Chapter 2.6.1 --- Amplification of KIAA0022 gene using PCR --- p.24 / Chapter 2.6.2 --- Purification of the PCR product --- p.25 / Chapter 2.6.3 --- Ligation --- p.25 / Chapter 2.6.4 --- One Shot® TOP 10 Chemical Transformation --- p.25 / Chapter 2.6.5 --- Small-scale preparation of the plasmid DNA --- p.26 / Chapter 2.6.6 --- Large-scale preparation of the plasmid DNA Table of Contents (continued) --- p.26 / Chapter 2.6.7 --- DNA sequencing of the full-length cDNA of KIAA0022 --- p.28 / Chapter 2.7 --- Northern Hybridization --- p.29 / Chapter 2.7.1 --- The Human multiple tissue Northern Blot --- p.29 / Chapter 2.7.2 --- Synthesis of the radiolabeled DNA probe --- p.29 / Chapter 2.7.3 --- Hybridization of the Northern blot --- p.30 / Chapter 2.8 --- Subcellular localization of KIAA0022 by tagging with green fluorescence protein (GFP) --- p.30 / Chapter 2.8.1 --- Amplification and purification of the KIAA0022 gene product --- p.30 / Chapter 2.8.2 --- Restriction enzymes digestion --- p.31 / Chapter 2.8.3 --- DNA ligation --- p.31 / Chapter 2.8.4 --- Preparation of the Escherichia coli competent cells for transformation --- p.31 / Chapter 2.8.5 --- Transformation of the plasmid DNA into competent Escherichia coli cells --- p.32 / Chapter 2.8.6 --- Small-scale preparation of the plasmid DNA --- p.32 / Chapter 2.8.7 --- Large-scale preparation of the plasmid DNA --- p.32 / Chapter 2.8.8 --- DNA sequencing of the cloned plasmid DNA --- p.33 / Chapter 2.8.9 --- Transfection --- p.33 / Chapter 2.8.10 --- Fluorescence microscopy examination --- p.33 / Chapter 2.9 --- Yeast two-hybrid screening assay --- p.34 / Chapter 2.9.1 --- "Cloning of the KIAA0022 gene into the yeast two-hybrid DNA-BD vector, pGBKT7" --- p.34 / Chapter 2.9.2 --- Small-scale transformation of pGBKT7-KIAA0022 plasmid --- p.34 / Chapter 2.9.2.1 --- Preparation of yeast competent cells --- p.34 / Chapter 2.9.2.2 --- Transformation of the pGBKT7-KIAA 0022 plasmid into the yeast strain PJ69-2A --- p.35 / Chapter 2.9.3 --- Screening a pretransformed library by yeast mating --- p.35 / Chapter 2.9.4 --- β -Galactosidase analysis - colony lift filter assay --- p.36 / Chapter 2.9.5 --- Analysis of yeast plasmid inserts using PCR and DNA sequencing --- p.37 / Chapter 2.9.5.1 --- PCR --- p.37 / Chapter 2.9.5.2 --- DNA sequencing --- p.37 / Chapter Chapter 3 --- Results / Chapter 3.1 --- Results of ESTs sequencing in normal counterpart and HCC libraries --- p.38 / Chapter 3.1.1 --- The sequencing results of the normal counterpart cDNA clones --- p.38 / Chapter 3.1.2 --- Sequencing results of the human liver cancer cDNA clones --- p.41 / Chapter 3.1.3 --- The accuracy of the automated sequencing technique --- p.41 / Chapter 3.1.4 --- Catalogue of normal counterpart ESTs --- p.45 / Chapter 3.1.5 --- Catalogue of liver cancer ESTs --- p.47 / Chapter 3.2 --- Identification of genes differentially expressed in HCC using in silico method --- p.115 / Chapter 3.3 --- Sequence analysis of KIAA0022 --- p.121 / Chapter 3.3.1 --- Structural analysis of KIAA0022 --- p.121 / Chapter 3.3.2 --- Homology alignment --- p.122 / Chapter 3.4 --- Tissue distribution and expression profile of KIAA0022 using Northern blot analysis --- p.132 / Chapter 3.5 --- Subcellular localization of the KIAA0022 tagging by green fluorescence protein --- p.134 / Chapter 3.6 --- Yeast two-hybrid screening assay --- p.136 / Chapter Chapter 4 --- Discussion / Chapter 4.1 --- Large-scale partial cDNA sequencing --- p.138 / Chapter 4.2 --- Characterization of ESTs --- p.139 / Chapter 4.3 --- Identification of genes differentially expressed in liver cancer using Poisson probability --- p.143 / Chapter 4.4 --- Characterization of KIAA0022 --- p.154 / Reference --- p.157 / Appendix --- p.170
| Identifer | oai:union.ndltd.org:cuhk.edu.hk/oai:cuhk-dr:cuhk_324081 |
| Date | January 2002 |
| Contributors | Au, Chi Chuen., 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, x, 173 leaves : ill. ; 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/) |
Page generated in 0.0026 seconds