by Liu Chun Ki, Kevin. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1999. / Includes bibliographical references (leaves 126-142). / Abstracts in English and Chinese. / Abstract --- p.i / Acknowledgements --- p.ii / Table of contents --- p.iii / List of figures --- p.viii / List of tables --- p.x / List of abbreviations --- p.xi / Chapter Chapter 1 --- Introduction Epstein-Barr Virus / Chapter 1.1 --- History --- p.1 / Chapter 1.2 --- Classification --- p.2 / Chapter 1.3 --- Virus and genome structure --- p.3 / Chapter 1.4 --- Epidemiology --- p.6 / Chapter 1.4.1 --- Prevalence of infection --- p.6 / Chapter 1.4.2 --- Modes of transmission --- p.7 / Chapter 1.5 --- Pathogenesis of EBV --- p.7 / Chapter 1.5.1 --- "Adsorption, penetration and dissemination" --- p.7 / Chapter 1.5.2 --- Lytic infection cycle --- p.8 / Chapter 1.5.3 --- Latent infection cycle --- p.9 / Chapter 1.5.4 --- Functions of the EBV-specific proteins associated with latent infection cycle proteins --- p.10 / Chapter 1.5.4.1 --- EBNA1 --- p.10 / Chapter 1.5.4.2 --- EBNA2 --- p.11 / Chapter 1.5.4.3 --- "EBNA 3A, 3B and 3C" --- p.11 / Chapter 1.5.4.4 --- EBNA LP --- p.12 / Chapter 1.5.4.5 --- LMP1 --- p.13 / Chapter 1.5.4.6 --- Characteristics of EBV LMP 2 gene --- p.14 / Chapter 1.5.4.7 --- Functions of LMP 2A --- p.15 / Chapter 1.5.4.8 --- Functions of LMP 2B --- p.18 / Chapter 1.6 --- Clinical significance of EBV --- p.20 / Chapter 1.6.1 --- Infectious mononucleosis (IM) --- p.20 / Chapter 1.6.2 --- Burkitt's lymphoma (BL) --- p.20 / Chapter 1.6.3 --- Nasopharyngeal carcinoma (NPC) --- p.21 / Chapter 1.6.4 --- Hodgkin's lymphoma (HL) --- p.21 / Chapter 1.7 --- Immune response to EBV infection --- p.22 / Chapter 1.7.1 --- Humoral immune response --- p.22 / Chapter 1.7.2 --- Cellular immune response --- p.22 / Chapter 1.8 --- Diagnosis of EBV infection --- p.26 / Chapter 1.9 --- Treatment and prevention --- p.27 / Chapter 1.10 --- Nasopharygneal Carcinoma (NPC) --- p.28 / Chapter 1.10.1 --- Epidemiology --- p.28 / Chapter 1.10.2 --- Etiology --- p.28 / Chapter 1.10.2.1 --- Environmental factor associated with NPC --- p.30 / Chapter 1.10.2.2 --- Genetic factors associated with NPC --- p.31 / Chapter 1.10.2.3 --- Association of NPC and EBV --- p.31 / Chapter 1.10.3 --- Diagnosis ofNPC --- p.32 / Chapter 1.10.4 --- Treatment --- p.33 / Chapter 1.11 --- Objective of the project --- p.34 / Chapter Chapter 2 --- Materials and Methods / Chapter 2.1 --- EBV-containing cell cultures --- p.35 / Chapter 2.2 --- Extraction of total RNA --- p.36 / Chapter 2.2.1 --- Cell lysis --- p.36 / Chapter 2.2.2 --- Protein digestion --- p.36 / Chapter 2.2.3 --- DNA digestion --- p.37 / Chapter 2.2.4 --- Elution of total RNA --- p.37 / Chapter 2.2.5 --- Purity and electrophoresis analysis of total RNA --- p.38 / Chapter 2.3 --- First strand cDNA synthesis --- p.38 / Chapter 2.4 --- PCR amplification of LMP 2 cDNA --- p.39 / Chapter 2.5 --- Isolation of the PCR amplified LMP 2 cDNA --- p.40 / Chapter 2.6 --- Purification of the PCR amplified LMP 2 cDNA --- p.41 / Chapter 2.7 --- Confirmation of the PCR amplified cDNA --- p.42 / Chapter 2.7.1 --- Nested PCR --- p.42 / Chapter 2.7.2 --- Restriction enzyme digestion --- p.44 / Chapter 2.8 --- Ligation of insert LMP 2 cDNA with vector --- p.45 / Chapter 2.9 --- Transformation of competent cells JM109 --- p.45 / Chapter 2.10 --- Screening of the recombinant clones --- p.47 / Chapter 2.11 --- Small scale purification of plasmid DNA --- p.47 / Chapter 2.12 --- Determination of the size of the insert DNA --- p.48 / Chapter 2.13 --- DNA sequencing --- p.48 / Chapter 2.13.1 --- The cycle sequencing reaction --- p.48 / Chapter 2.13.2 --- Preparation of the acrylamide gel and TBE buffer --- p.51 / Chapter 2.13.3 --- Running conditions of the electrophoresis --- p.52 / Chapter 2.13.4 --- "Processing, editing and exporting the sequences" --- p.52 / Chapter 2.14 --- Data analysis --- p.53 / Chapter 2.14.1 --- Sequence analysis --- p.53 / Chapter 2.14.2 --- Amino acid analysis --- p.53 / Chapter 2.14.3 --- Protein secondary structure analysis --- p.53 / Chapter 2.14.4 --- Hydrophobicity analysis --- p.54 / Chapter 2.14.5 --- Isoelectric point analysis --- p.54 / Chapter Chapter 3 --- Results / Chapter 3.1 --- Cell Cultures --- p.55 / Chapter 3.2 --- Extraction of total RNA --- p.56 / Chapter 3.3 --- PCR amplification --- p.61 / Chapter 3.4 --- Isolation of PCR amplified LMP 2 cDNA --- p.62 / Chapter 3.5 --- Confirmation of the PCR amplified cDNA --- p.66 / Chapter 3.5.1 --- Nested PCR --- p.66 / Chapter 3.5.2 --- Restriction enzyme digestion --- p.71 / Chapter 3.6 --- Transformation and screening --- p.77 / Chapter 3.7 --- Extraction of plasmid DNA and its digestion with restriction enzyme --- p.78 / Chapter 3.8 --- DNA sequencing --- p.83 / Chapter 3.8.1 --- DNA sequence comparison --- p.84 / Chapter 3.9 --- Amino acid sequence homology --- p.89 / Chapter 3.9.1 --- Amino acid sequence comparison --- p.90 / Chapter 3.10 --- Hydrophobicity analysis --- p.92 / Chapter 3.10.1 --- Comparison of hydrophobicity of B95-8 derived LMP2 with GeneBank --- p.93 / Chapter 3.10.2 --- Comparison of hydrophobicity of CB 14022-derived LMP2 with GeneBank --- p.95 / Chapter 3.10.3 --- Comparison of hydrophobicity of Raji-derived LMP2 with GeneBank --- p.97 / Chapter 3.11 --- Protein secondary structure analysis --- p.100 / Chapter 3.11.1 --- Comparison of secondary structure of B95-8-derived LMP2 with GeneBank --- p.100 / Chapter 3.11.2 --- Comparison of secondary structure of CB 14022-derived LMP2 with GeneBank --- p.100 / Chapter 3.11.3 --- Comparison of secondary structure of Raji-derived LMP2 with GeneBank --- p.101 / Chapter 3.12 --- Isoelectric point analysis --- p.103 / Chapter Chapter 4 --- Discussions / Chapter 4.1 --- Overall strategy for the cloning and sequencing of EBV LMP 2 gene --- p.106 / Chapter 4.2 --- Implications of the results obtained in sequencing --- p.107 / Chapter 4.3 --- Results interpretation --- p.108 / Chapter 4.3.1 --- Cell culture --- p.108 / Chapter 4.3.2 --- Extraction of total RNA --- p.108 / Chapter 4.3.3 --- PCR amplification --- p.109 / Chapter 4.3.4 --- Confirmation of the PCR amplified cDNAs using nested PCR --- p.109 / Chapter 4.3.5 --- Confirmation of the PCR amplified cDNAs using restriction enzyme digestion --- p.110 / Chapter 4.3.6 --- Ligation of EBV LMP 2 cDNA to pGEM-T Easy Vector --- p.111 / Chapter 4.3.7 --- Transformation and screening --- p.114 / Chapter 4.3.8 --- Extraction of plasmid DNA and digestion with restriction enzyme --- p.115 / Chapter 4.4 --- DNA sequencing and sequence homology --- p.116 / Chapter 4.5 --- Amino acid sequence homology --- p.117 / Chapter 4.6 --- Hydrophobicity analysis --- p.119 / Chapter 4.7 --- Protein secondary structure analysis --- p.120 / Chapter 4.8 --- Isoelectric point analysis --- p.122 / Chapter 4.9 --- Summary of results --- p.122 / Chapter 4.10 --- Conclusions --- p.124 / References --- p.126
| Identifer | oai:union.ndltd.org:cuhk.edu.hk/oai:cuhk-dr:cuhk_322858 |
| Date | January 1999 |
| Contributors | Liu, Chun Ki Kevin., Chinese University of Hong Kong Graduate School. Division of Microbiology. |
| Source Sets | The Chinese University of Hong Kong |
| Language | English, Chinese |
| Detected Language | English |
| Type | Text, bibliography |
| Format | print, xii, 142 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/) |
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