Student Number : 9901196J - MSc dissertation - School of Pathology - Faculty of Science / Fragile sites are highly unstable regions of the genome, which have a tendency to form gaps and breaks in metaphase chromosomes under replication stress conditions. There are many common fragile sites in the human genome and exposure to carcinogens may affect several genes localised in fragile sites within a single cell, which could lead to activation of oncogenes and inactivation of tumour-suppressor genes simultaneously. FRA3B on chromosome 3 and FRA16D on chromosome 16 are the two most commonly expressed fragile sites and contain the FHIT and WWOX genes respectively. These genes are tumour suppressor genes and are inactivated in a number of different ways. Carcinogens found in cigarette smoke have been found to increase fragile site expression and could alter the integrity of theses genes in active smokers. Ten healthy non-smoking (control) individuals and twenty active smokers were recruited for the purpose of this study. Fluorescence in situ hybridisation was performed with probes spanning spanning the FHIT gene and RT-PCR was performed to assess both FHIT and WWOX expression. No significant difference in breaks at fragile sites was observed between controls and active smokers in the FISH experiments. In addition, no aberrant transcripts were detected for either FHIT or WWOX with RT-PCR. Although the sampling group was limited and heterogenous, no increase in the expression of breaks at fragile sites was seen in active smokers in the present study.

by Law Hon Man. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1993. / Includes bibliographical references (leaves 67-70). / Chapter 1 --- Introduction --- p.1 / Chapter 2 --- Automated Chromosome Classification --- p.4 / Chapter 2.1 --- Procedures in Chromosome Classification --- p.6 / Chapter 2.2 --- Sample Preparation --- p.7 / Chapter 2.3 --- Low Level Processing and Measurement --- p.9 / Chapter 2.4 --- Feature Extraction --- p.11 / Chapter 2.5 --- Classification --- p.15 / Chapter 3 --- Inference of Markov Networks by Dynamic Programming --- p.17 / Chapter 3.1 --- Markov Networks --- p.18 / Chapter 3.2 --- String-to-String Correction --- p.19 / Chapter 3.3 --- String-to-Network Alignment --- p.21 / Chapter 3.4 --- Forced Landmarks in String-to-Network Alignment --- p.31 / Chapter 4 --- Landmark Finding in Markov Networks --- p.34 / Chapter 4.1 --- Landmark Finding without a priori Knowledge --- p.34 / Chapter 4.2 --- Chromosome Profile Processing --- p.37 / Chapter 4.3 --- Analysis of Chromosome Networks --- p.39 / Chapter 4.4 --- Classification Results --- p.45 / Chapter 5 --- Speech Recognition using Inferred Markov Networks --- p.48 / Chapter 5.1 --- Linear Predictive Analysis --- p.48 / Chapter 5.2 --- TIMIT Speech Database --- p.50 / Chapter 5.3 --- Feature Extraction --- p.51 / Chapter 5.4 --- Empirical Landmarks in Speech Networks --- p.52 / Chapter 5.5 --- Classification Results --- p.55 / Chapter 6 --- Conclusion --- p.57 / Chapter 6.1 --- Suggested Improvements --- p.57 / Chapter 6.2 --- Concluding remarks --- p.61 / Appendix A --- p.63 / Reference --- p.67

Orientador: Edson Luiz Furtado / Coorientador: Celso Luiz Marino / Banca: Esteban Roberto González / Banca: Mario Tomazello Filho / Resumo: A poliploidia tem sido um importante mecanismo de destaque na história evolutiva das plantas e outros eucariotos. A ocorrência disseminada de poliplóides na natureza sugere que possa existir uma função para a sobrevivência e colonização em novos ambientes. No entanto, até agora foram poucos os estudos que exploraram esses fenômenos em espécies de Eucalyptus. Neste trabalho foram caracterizadas e comparadas 25 plantas tetraplóides, 25 diplóides pós contato com colchicina e 12 diplóides sem contato com colchicina de Eucalyptus dunnii em casa de vegetação. Foram medidas as alterações morfológicas por um período de seis meses. Concluiu-se que o número de ramos, dimensões de folhas (comprimento, largura e pecíolo), frequência de estômatos e comprimento de fibras mostraram diferenças significativas entre plantas diplóides e tetraplóides. Embora o número de galhos tenha diminuído em plantas tetraplóides, as plantas testadas não apresentaram diferenças significativas na altura e diâmetro. Estes resultados demonstraram que existem diferenças morfológicas entre plantas diplóides e tetraplóides de Eucalyptus dunnii e que estas diferenças podem ser usadas para facilitar a identificação de plantas tetraplóides recentemente produzidas. / Abstract: Polyploidy has been an important mechanism in the plant evolution and other eukaryotes. The widespread occurrence of polyploid in nature suggest that there may be a function for survival and colonization of new environments. However, so far there are a very few studies that have explored these phenomena in Eucalyptus species. In this work were characterized and compared 25 tetraploid plants, 25 diploid after contact with colchicine and 12 diploid without contact with colchicine of Eucalyptus dunnii under greenhouse conditions. Morphological changes were measured for a period of six months. It was concluded that the number of branches, leaf area (length, width and petiole), frequency of stomata and fibre length showed significant differences between diploid and tetraploid plants. Although the number of branches has decreased in tetraploid plants and the plants tested showed no significant differences in height. These results demonstrated that there are morphological differences between diploid and tetraploid Eucalyptus and that these differences may be used to facilitate identification of newly produced tetraploid plants / Mestre

Oligoryzomys é o gênero mais especioso da tribo Oryzomyini e está amplamente distribuído na região Neotropical. O objetivo deste trabalho é contribuir para a citotaxonomia e investigar a evolução cromossômica no gênero. Foram analisados 117 exemplares pertencentes às espécies: O. flavescens (2n=64-66, NF=66), O. fornesi (2n=62, NF=64), O. microtis (2n=64, NF=64), O. moojeni (2n=70, NF=72), O. nigripes (2n=62, NF=78-82), O. stramineus (2n=52, NF=68) e Oligoryzomys sp. A (2n=70, NF=72). As seis primeiras possuem cariótipos espécie-específicos e, dessa forma, reiteramos a importância da informação citogenética para a citotaxonomia. A pintura cromossômica comparativa (Zoo-FISH) com sondas de O. moojeni revelou hibridação em 29 segmentos autossômicos em O. fornesi; 30 em O. microtis; 31 em O. nigripes; e 32 em O. rupestris e Oligoryzomys sp. 2. Os resultados mostraram uma extensa reorganização genômica na evolução cromossômica do gênero, decorrente de fissões, fusões em tandem, rearranjos Robertsonianos e perda/inativação, surgimento ou reposicionamento de centrômero. / Oligoryzomys is the most specious genus within the tribe Oryzomyini and it is distributed throughout Neotropical region. This work aims to contribute to citotaxonomy and to investigate the chromosomal evolution of the genus. A total of 117 individuals were cytogenetically analysed, and they belong to the species: O. flavescens (2n=64-66, FN=66), O. fornesi (2n=62, FN=64), O. microtis (2n=64, FN=64), O. moojeni (2n=70, FN=72), O. nigripes (2n=62, FN=78-82), O. stramineus (2n=52, FN=68), and Oligoryzomys sp. A (2n=70, FN=72). The first six species possess species-specific karyotypes, and therefore we emphasize the importance of cytogenetic studies for citotaxonomy. Comparative chromosome painting (Zoo-FISH) with O. moojeni probes hybridized to 29 segments on metaphases of O. fornesi, 30 on O. microtis, 31 on O. nigripes, and 32 on O. rupestris and Oligoryzomys sp. 2. The results showed an extensive genomic reshuffling, due to fissions, tandem and Robertsonian fusions, loss/inactivation or repositioning of centromeres.

Leung Chin-lung. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2003. / Includes bibliographical references (leaves 133-164). / Abstracts in English and Chinese. / Abstract --- p.iv / 摘要 --- p.vi / List of abbreviation --- p.viii / Chapter Chapter 1 --- Hepatocellular Carcinoma --- p.1 / Chapter 1.1 --- A Health Burden --- p.1 / Chapter 1.2 --- Pathology --- p.3 / Chapter 1.3 --- Epidemiology --- p.7 / Chapter 1.3.1 --- Global HCC distribution --- p.7 / Chapter 1.3.2 --- Age and Gender --- p.10 / Chapter 1.4 --- Risk Factors of HCC --- p.12 / Chapter 1.4.1 --- Hepatitis B virus (HBV) --- p.13 / Chapter 1.4.1.1 --- Chronic HBV infection --- p.13 / Chapter 1.4.1.2 --- Role of HBV in hepatocarcinogenesis --- p.16 / Chapter 1.4.1.2 a) --- Direct Oncogenesis --- p.16 / Chapter 1.4.1.2 b) --- Indirect Oncogenesis --- p.17 / Chapter 1.4.2 --- Hepatitis C virus (HCV) --- p.23 / Chapter 1.4.2.1 --- Chronic HCV infection --- p.23 / Chapter 1.4.2.2 --- Role of HCV in hepatocarcinogenesis --- p.23 / Chapter 1.4.3 --- Chemicals as liver carcinogens --- p.27 / Chapter 1.4.3.1 --- Aflatoxin Bi (AFB1) --- p.28 / Chapter 1.4.3.2 --- Vinyl chloride --- p.29 / Chapter 1.4.3.3 --- Alcoholic beverages --- p.29 / Chapter 1.4.4 --- Inborn Errors in Metabolisms --- p.30 / Chapter 1.4.4.1 --- Hereditary tyrosinemia --- p.30 / Chapter 1.4.4.2 --- Hereditary haemochromatosis --- p.30 / Chapter 1.4.4.3 --- α1-antitrypsin deficiency --- p.31 / Chapter 1.4.5 --- Liver lesions --- p.32 / Chapter 1.5 --- Genetic alterations in HCC --- p.33 / Chapter Chapter 2 --- Rationale of the study --- p.39 / Chapter Chapter 3 --- LOH study on 8p in HCC --- p.48 / Chapter 3.1 --- Introduction --- p.48 / Chapter 3.1.1 --- "Knudson's ""two-hit"" model and LOH" --- p.48 / Chapter 3.1.2 --- Microsatellite DNA and LOH study --- p.49 / Chapter 3.2 --- Materials and Methods --- p.51 / Chapter 3.2.1 --- Patients and Specimens --- p.51 / Chapter 3.2.1.1 --- Genomic DNA extraction from liver tissues --- p.53 / Chapter 3.2.1.2 --- Genomic DNA extraction from buffy coat --- p.55 / Chapter 3.3 --- LOH study on 8p in HCC --- p.57 / Chapter 3.3.1 --- Microsatellite markers --- p.57 / Chapter 3.3.2 --- 5-end labeling --- p.60 / Chapter 3.3.3 --- Amplification of microsatellite DNA --- p.60 / Chapter 3.3.4 --- Denaturing polyacrylamide gel electrophoresis --- p.61 / Chapter 3.3.5 --- Detection of LOH --- p.62 / Chapter 3.4 --- Results --- p.63 / Chapter 3.4.1 --- LOH status of 52 HCC cases --- p.63 / Chapter 3.4.2 --- Clinicopathological correlation --- p.67 / Chapter 3.4.3 --- Delineation of common deletion region (CDR) --- p.67 / Chapter 3.4.4 --- Common deletion region of interest --- p.77 / Chapter Chapter 4 --- Study on LZTS1 --- p.83 / Chapter 4.1 --- Introduction 一 LZTS1 --- p.83 / Chapter 4.2 --- Mutation analysis of LZTS1 in HCC --- p.87 / Chapter 4.2.1 --- Materials and Methods --- p.87 / Chapter 4.2.1.1 --- Patients and HCC cell lines --- p.87 / Chapter 4.2.1.2 --- Genomic DNA extraction from HCC cell lines --- p.87 / Chapter 4.2.1.3 --- Amplification of exons of LZTS1 --- p.89 / Chapter 4.2.1.3a) --- Primer pairs --- p.89 / Chapter 4.2.1.3b) --- PCR conditions --- p.90 / Chapter 4.2.1.4 --- Purification of PCR products --- p.93 / Chapter 4.2.1.5 --- Cycle sequencing reaction --- p.94 / Chapter 4.2.1.6 --- Purification of cycle sequencing reaction product --- p.94 / Chapter 4.2.1.7 --- Sequence analysis by automated sequencer --- p.95 / Chapter 4.2.1.8 --- Search for sequence variants of LZTS1 --- p.96 / Chapter 4.2.2 --- Results --- p.97 / Chapter 4.3 --- Expression analysis of LZTS1 in HCC with preliminary results --- p.103 / Chapter 4.3.1 --- Materials and Methods --- p.103 / Chapter 4.3.1.1 --- Patients and Specimens --- p.103 / Chapter 4.3.1.2 --- Total RNA extraction --- p.103 / Chapter 4.3.1.3 --- Reverse transcription --- p.104 / Chapter 4.3.1.4 --- Semi-quantitative PCR --- p.105 / Chapter 4.3.1.4a) --- Primer pairs --- p.105 / Chapter 4.3.1.4b) --- PCR conditions --- p.106 / Chapter 4.3.2 --- Results --- p.109 / Chapter Chapter 5 --- Discussion --- p.111 / Chapter 5.1 --- LOH study on 8p in HCC --- p.111 / Chapter 5.2 --- Study on LZTS1 in HCC --- p.125 / Chapter 5.2.1 --- Mutation analysis of LZTS1 --- p.125 / Chapter 5.2.2 --- Expression analysis of LZTS1 --- p.129 / Chapter 5.3 --- Future Study --- p.132 / References --- p.133

by Angela Bik-Yu Hui. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1997. / Includes bibliographical references (leaves 119-133). / Acknowledgements / Table of Contents / List of Tables / List of Figures / Abstract / Chapter CHAPTER 1 --- LITERATURE REVIEW --- p.1 / Chapter I. --- Nasopharyngeal carcinoma --- p.1 / Chapter II. --- Etiology of NPC --- p.3 / Chapter II a. --- Geographical & Environmental factors --- p.3 / Chapter II b. --- Epstein-Barr virus Infection --- p.5 / Chapter II c. --- Genetic Factors --- p.8 / Chapter III. --- Cytogenetic Studies of NPC --- p.10 / Chapter III a. --- Traditional Cytogenetics --- p.10 / Chapter III b. --- Previous cytogenetic findings of NPC --- p.12 / Chapter III.c. --- Fluorescence in-situ hybridization --- p.15 / Chapter III.d. --- The new NPC cell line: Cell-666 --- p.18 / Chapter IV. --- Molecular Genetic Studies in NPC --- p.19 / Chapter IV a. --- Oncogenes --- p.20 / Chapter IV b. --- Tumor suppresser genes (TSGs) --- p.22 / Chapter IV c. --- Loss of Heterozygosity Studies --- p.29 / Chapter IV d. --- LOH on Chromosome 11 --- p.32 / Chapter IV e. --- ATM Gene --- p.35 / Chapter CHAPTER 2 --- OBJECTIVE OF STUDY --- p.38 / Chapter CHAPTER 3 --- MATERIALS AND METHODS --- p.41 / Chapter I： --- Study of loss of heterozygosity on chromosome 11 --- p.41 / Chapter I a. --- Patients and Specimens --- p.41 / Chapter I.b. --- DNA extraction --- p.45 / Chapter I c. --- Microsatellite Polymorphism Analysis --- p.47 / Chapter I d. --- Multiplex PCR analysis --- p.52 / Chapter II. --- Cytogenetic Studies --- p.54 / Chapter II a. --- Culture of cell-666 --- p.54 / Chapter II b. --- Cytogenetic Analysis --- p.56 / Chapter II c. --- Fluorescence in-situ hybridization (FISH) --- p.58 / Chapter II d. --- FISH analysis of other NPC cell lines) --- p.62 / Chapter CHAPTER 4 --- RESULTS --- p.63 / Chapter I: --- Study of loss of heterozygosity on chromosome 11 --- p.63 / Chapter I a. --- LOH analysis --- p.63 / Chapter I b. --- Regions with L OH --- p.73 / Chapter I c. --- Multiplex PCR analysis --- p.79 / Chapter II: --- Cytogenetic Study --- p.83 / Chapter II a. --- Cytogenetic analysis of cell-666 --- p.83 / Chapter II.b. --- Fluorescence in-situ Hybridization (FISH) --- p.91 / Chapter CHAPTER 5 --- DTSCUSSION --- p.102 / Chapter I. --- LOH of Chromosome 11 Studies --- p.102 / Chapter II. --- Comparison with LOH studies of other chromosomes --- p.110 / Chapter III. --- Cytogenetic Studies --- p.113 / REFERENCES --- p.119

It is incumbent upon cellular life to ensure the faithful transmission of genetic material from mother cell to daughter cell and from parent to progeny. However, cells are under constant threat of DNA damage from sources both endogenous and exogenous, such as the products of metabolism and genotoxic chemicals. Thus, cells have evolved multiple systems of repair to ensure genome integrity. The DNA double-strand break (DSB) is among the most lethal forms of DNA damage, and a critical pathway to resolve these lesions is homologous recombination (HR). During HR, information lost at the cut site of one locus is repaired when the damaged site locates a homologous sequence in the nucleus to use as template for repair. The process by which a cut chromosome finds its homolog is known as homology search, and, while the enzymatic steps of HR have been well studied in recent years, the coordination of cell biological events like HS in the context of the crowded nucleus has remained poorly understood. Recently, our laboratory and others have studied a phenomenon known as DNA damage-induced increased chromosomal mobility, in which chromosomal loci, both damaged and undamaged, explore larger areas of the nucleus after the formation of DSBs. The increase in the mobility of cut loci is known as local mobility, and the increase in mobility of undamaged loci in response to a break elsewhere in the nucleus is known as global mobility. Here, I report that the recombination machinery and the DNA damage checkpoint cooperate in order to regulate global mobility of chromosomes following DSB formation. The RecA-like recombinase Rad51 is required for global mobility, and exerts its effect at single-stranded DNA (ssDNA), but its canonical homology search and strand exchange functions are not required. I find that Rad51 is ultimately required to displace Rad52, which is revealed to be an inhibitor of mobility when bound to ssDNA in the absence of Rad51. Thus, recombination factors can serve as DNA damage sensors, and relay information to the checkpoint apparatus in order to govern the initiation of increased mobility after DSB formation. I have also studied how the baseline confinement of loci is established, and assessed the contributions of several genes involved in repair to increased mobility. These observations offer novel insight into previously unappreciated regulatory functions performed by the recombination machinery, and demonstrate how the progression of DNA repair pathways influences nuclear organization.

The autoimmune diseases embody a diverse range of common human conditions that are caused by a loss of self-tolerance in the host immune system to a specific organ or tissue type. Approximately 5% of the general population are affected by autoimmune diseases which include type 1 diabetes (T1D), rheumatoid arthritis (RA) and Graves disease (GD). The majority of the autoimmune diseases are multifactorial in origin, brought about by a combination of both environmental and genetic factors. Numerous susceptibility loci have been identified for each autoimmune disease and a number of these loci have been shown to be shared amongst the autoimmune diseases. The fine-mapping of susceptibility loci to the underlying disease genes remains the current challenge facing complex disease genetics. This project aimed to further characterise the autoimmune disease susceptibility locus IDDM6 on chromosome 18q12-21. This was achieved by using a comparative mapping approach that incorporates the study of genetic association in human autoimmune disease alongside the consomic mapping of the orthologous region in the non-obese diabetic (NOD) mouse model of autoimmunity. Deleted in colorectal carcinomas (DCC) provided a strong candidate gene at IDDM6 and the resident R201G polymorphism was identified as a functional candidate A potential mechanism for the R201G polymorphism involvement in T1D aetiology was identified where the polymorphism may affect the ability of DCC to induce apoptosis in vitro. However, no evidence for R201G association could be detected in autoimmune disease case-control datasets from the New Zealand (NZ) population (T1D n = 428, RA n = 730, autoimmune thyroid disease n = 192 (AITD); versus n = 1246 healthy controls). In addition, no evidence for R201G involvement in T1D could be provided in a transmission disequilibrium test (TDT) incorporating 382 affected sib-pair families (54.2% transmission; P = 0.15). Significant association of R201G with GD was detected in a United Kingdom (UK) dataset (P = 0.002) from the Newcastle population (423 cases vs. 393 controls) but this was not replicated in an additional dataset from the UK Birmingham population (731 cases vs 668 controls; P = 0.81). It was concluded that the R201G polymorphism may encode susceptibility to GD but is unlikely to be the sole aetiological variant that accounts for the linkage previously observed at IDDM6 in autoimmune disease. To further investigate DCC as a positional candidate at IDDM6, five SNPs were selected from a 100 kb window surrounding a DCC-resident microsatellite that had previously been associated with T1D, called "88,21". The five SNPs were genotyped in the NZ T1D dataset, and the ascertainment of estimated haplotypes in this dataset revealed association of a rare haplotype with T1D, called haplotype H (3.31% cases vs 1.17% controls; P = 0.0044), in addition to global association of all haplotypes (P = 0.018). Haplotype H was also associated in an independent case-control dataset from the UK comprised of 400 T1D subjects and 443 healthy controls (P = 0.038). Maximum support for association of haplotype H was extended when both the UK and NZ T1D datasets were combined (P = 0.0017). Association of haplotype H could not be verified in a family-based test for association using the 382 UK T1D families (P = 0.40). However, the inclusion of the DCC SNPs in a TDT analysis of the published DCC-resident microsatellites "88,21" and "55,26", that had been used to identify IDDM6, extends support for the previously-associated 2-10 haplotype (2-10 refers to the published allele nomenclature at "88,21" and "55,26" respectively; 2-10-haplotype A; 59.6% T; P = 0.0058). There was no evidence for association of the five SNPs with RA or AITD when using either individual SNP analyses or estimated haplotypes in the NZ datasets. A similar lack of association was reported for the UK Newcastle GD dataset. Taken together, these data further support DCC, or a nearby gene, as conferring susceptibility to T1D. The human genetic data that supports IDDM6 involvement in autoimmune disease is further strengthened by consomic mapping of the orthologous region in mouse, using the non-obese diabetic mouse (NOD) model of autoimmune disease. In this thesis, the first evidence for a diabetes and thyroiditis susceptibility locus on mouse chromosome 18 is presented, which have been designated Idd21 and Sat1 respectively. This was achieved by using a chromosome-replacement strain with chromosome 18 derived from the diabetes-resistant Biozzi ABH strain on a diabetes-susceptible NOD genome, called NOD.ABH[Chr�⁸]. Mouse chromosome 18 contains orthology to both IDDM6 and the rat diabetes-susceptibility locus Iddm3. The NOD.ABH[Chr�⁸] mice showed a dramatic and significant reduction in diabetes incidence (30% of females were affected by 7 months of age versus 85% in NOD; P <0.0001) and that of thyroiditis (15.5% at 12 months compared to 37.4% in NOD; P <0.002). The comparative mapping of the chromosome 18 autoimmune susceptibility locus IDDM6 in human and mouse presented in this thesis provides further support for this locus. This research also clearly defines the next steps required to fine-map IDDM6 to the underlying disease genes, especially in regard to the DCC gene.

Errata pasted on front fly-leaf. Bibliography: leaves 139-173. This thesis identifies DNA structures and genes involved in the process of homologous chromosome pairing in allohexaploid bread wheat (Triticum aestivum). In addition to studying late replicating DNA, a speculative model on the action of the pairing genes in allohexaploid wheat and the putative function of the AWWM5 gene is discussed.

No description available.