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by Nellie Yuk Fei Chung., Thesis (M.Phil.)--Chinese University of Hong Kong, 1998., Includes bibliographical references (leaves 111-120)., also in Chinese., p.i, List of Abbreviations --- p.iii, Acknowledgements --- p.iv, Publication of this study --- p.vi, Table of Contents --- p.vii, Chapter Chapter1: --- Introduction --- p.1, Chapter 1.1 --- Multiple Myeloma --- p.2, Chapter 1.2 --- The Problem --- p.2, Chapter Chapter2: --- Literature Review --- p.5, Chapter 2.1 --- Molecular Genetics of Multiple Myeloma --- p.6, Chapter 2.1.1 --- Cytogenetics --- p.6, Chapter 2.2 --- Alterations of Proto-Oncogenes --- p.9, Chapter 2.2.1 --- c-myc --- p.9, Chapter 2.2.2 --- Ras --- p.10, Chapter 2.2.3 --- Bcl-2 and Related Protein --- p.10, Chapter 2.3 --- Alteration of Tumor-Suppressor genes --- p.11, Chapter 2.3.1 --- p53 Gene Mutations --- p.11, Chapter 2.3.2 --- Retinoblastoma (Rb) Gene --- p.11, Chapter 2.3.3 --- p16 and p15 Genes --- p.13, Chapter Chapter3: --- DNA Methylation and Cancers --- p.14, Chapter 3.1 --- Role of DNA Methylation --- p.15, Chapter 3.2 --- CpG Islands --- p.15, Chapter 3.3 --- Abnormalities of DNA Methylation in Neoplasia --- p.16, Chapter 3.3.1 --- DNA Hypomethylation in Cancer --- p.16, Chapter 3.3.2 --- DNA Methyltransferase Activity in Cancer --- p.17, Chapter 3.4 --- Regional DNA Hypermethylation in Cancer --- p.17, Chapter 3.4.1 --- p16 and p15 Genes in Solid Tumors --- p.18, Chapter 3.4.2 --- The p16 and p15 Genes in Leukemia and other Hematopoietic Malignancies --- p.19, Chapter 3.4.3 --- Retinoblastoma Gene --- p.20, Chapter 3.5 --- Mechanism Underlying the DNA Methylation Changes --- p.21, Chapter Chapter4: --- Background of Study --- p.23, Chapter 4.1 --- Background of Study --- p.24, Chapter 4.2 --- Project Objectives --- p.27, Chapter Chapter5: --- Materials and Methods --- p.29, Chapter 5.1 --- Patients Samples --- p.30, Chapter 5.2 --- Normal Controls --- p.30, Chapter 5.3 --- Storage of the Samples --- p.32, Chapter 5.4 --- Materials --- p.32, Chapter 5.4.1 --- Chemicals --- p.32, Chapter 5.4.2 --- Primers --- p.33, Chapter 5.4.3 --- Enzymes --- p.35, Chapter 5.5 --- Methods --- p.35, Chapter 5.5.1 --- Cloning of p16 and p15 Exon 1 Probes for Southern Analysis --- p.35, Chapter 5.5.1.1 --- PCR Amplification of p16 and p15 exon1 Probes from Normal Blood DNA --- p.35, Chapter 5.5.1.2 --- Recovery and Purification of p16 and p15 Exon 1 DNA Fragment --- p.36, Chapter 5.5.1.3 --- Ligation --- p.37, Chapter 5.5.1.4 --- Transformation --- p.37, Chapter 5.5.1.5 --- Plating --- p.38, Chapter 5.5.1.6 --- Screening of Recombinant Plasmid --- p.38, Chapter 5.5.1.7 --- Confirmation of Cloned DNA by Sequencing --- p.42, Chapter 5.5.2 --- DNA Extraction and Purification --- p.45, Chapter 5.5.2.1 --- DNA Extraction from Bone Marrow Aspirate and Peripheral Blood --- p.45, Chapter 5.5.2.2 --- Isolation of Plasmid DNA from Transformant Cutures --- p.46, Chapter 5.5.2.3 --- Qualification and Quantification of DNA --- p.49, Chapter 5.5.3 --- Detection of Hypermethylation by Southern Analysis --- p.50, Chapter 5.5.3.1 --- Restriction Enzyme Digestion --- p.50, Chapter 5.5.3.2 --- Agarose Gel Electrophoresis --- p.51, Chapter 5.5.3.3 --- Southern Transfer --- p.51, Chapter 5.5.3.4 --- Membrane Fixation --- p.51, Chapter 5.5.3.5 --- Recovery and Purification of p16 and p15 Exon 1 Probes from Plasmid --- p.52, Chapter 5.5.3.6 --- Probe Labeling --- p.54, Chapter 5.5.3.7 --- Purification of Radioactive labeled DNA --- p.54, Chapter 5.5.3.8 --- Southern Hybridization --- p.55, Chapter 5.5.3.9 --- Post Hybridization --- p.55, Chapter 5.5.3.10 --- Autoradiography --- p.56, Chapter 5.5.4 --- Polymerase Chain Reaction-Single Strand Conformational Polymorphism Analysis (PCR-SSCP) --- p.56, Chapter 5.5.4.1 --- 5'- end Radioactive Labeling of Primer --- p.56, Chapter 5.5.4.2 --- Amplification of Target Sequence by PCR --- p.57, Chapter 5.5.4.3 --- Non-denaturing Polyacrylamide Gel Electrophresis --- p.57, Chapter 5.5.4.4 --- Direct DNA Sequence of PCR Products --- p.58, Chapter 5.5.5 --- Prevention of Overall Contamination in PCR --- p.60, Chapter 5.5.6 --- "Sensitivity, Specificity Controls" --- p.62, Chapter Chapter6: --- Results --- p.64, Chapter 6.1 --- Patient Characteristics --- p.65, Chapter 6.1.1 --- General Patient Characteristics --- p.65, Chapter 6.1.2 --- Clinical and Laboratory Features --- p.65, Chapter 6.2 --- Southern Blot Analysis of p16/p15 and Rb --- p.79, Chapter 6.2.1 --- Absence of Deletions or hypermethylationin Normal Controls --- p.79, Chapter 6.2.2 --- Absence of Homozygous Deletions or Mutationsin p16/15 and Rb among all MM Patients --- p.79, Chapter 6.2.3 --- Hypermethylation of p16 --- p.89, Chapter 6.2.4 --- Hypermethylation of p15 --- p.92, Chapter 6.3 --- Hypermethylation of p16/p15 and Clinico-pathologic Correlation --- p.94, Chapter Chapter7: --- Discussion --- p.97, Chapter 7.1 --- "Absence of Homozygous Deletions, Gene Rearrangements and Mutations in p16/p15 and Rb" --- p.98, Chapter 7.2 --- Hypermethylation of p16/p15-An Alternative Way for Gene Inactivation --- p.100, Chapter 7.2.1 --- Methylation of p15 Gene --- p.101, Chapter 7.2.2 --- Methylation of 5'-CpG Island of p16/p15 and Lack of Gene Expression --- p.102, Chapter 7.2.3 --- Comparison of Methylation Status of Primary Samples and Cell Lines in MM --- p.103, Chapter 7.2.4 --- Progressive Gene Inactivation by Random Methylation Errors --- p.104, Chapter 7.2.5 --- The Lack of Correlation of Tumor Contents Revealed by the Southern Analysis and Morphologic Assessment --- p.105, Chapter 7.3 --- Knudson's Two-hit Model of Tumorigenesis --- p.106, Chapter 7.4 --- Inverse Relationship of p16 and Rb --- p.107, Chapter 7.5 --- Implications of Our Findings --- p.109, Chapter 7.6 --- Future Studies --- p.109, References --- p.111, http://library.cuhk.edu.hk/record=b5889789, 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/)