published_or_final_version / abstract / Computer Science and Information Systems / Master / Master of Philosophy

published_or_final_version / Psychiatry / Doctoral / Doctor of Philosophy

<p>With easy access to technology to generate expressed sequence tags (ESTs), several groups have sequenced from thousands to several thousands of ESTs. These ESTs benefit from consolidation and organization to deliver significant biological value. A number of EST projects are underway to extract maximum value from fragmented EST resources by constructing gene indices, where all transcripts are partitioned into index classes such that transcripts are put into the same index class if they represent the same gene. Therefore a gene index should ideally represent a non-redundant set of transcripts. Indeed, most gene indices aim to reconstruct the gene complement of a genome and their technological developments are directed at achieving this goal. The South African National Bioinformatics Institute (SANBI), on the other hand, embarked on the development of the sequence alignment and consensus knowledgebase (STACK) database that focused on the detection and visualisation of transcript variation in the context of developmental and pathological states, using all publicly available ESTs. Preliminary work on the STACK project employed an approach of partitioning the EST data into arbitrarily chosen tissue categories as a means of reducing the EST sequences to manageable sizes for subsequent processing. The tissue partitioning provided the template material for developing error-checking tools to analyse the information embedded in the error-laden EST sequences. However, tissue partitioning increases redundancy in the sequence data because one gene can be expressed in multiple tissues, with the result that multiple tissue partitioned transcripts will correspond to the same gene.</p> <p><br /> Therefore, the sequence data represented by each tissue category had to be merged in order to obtain a comprehensive view of expressed transcript variation across all available tissues. The need to consolidate all EST information provided the impetus for developing a STACK human gene index, also referred to as a whole-body index. In this dissertation, I report on the development of a STACK human gene index represented by consensus transcripts where all constituent ESTs sample single or multiple tissues in order to provide the correct development and pathological context for investigating sequence variation. Furthermore, the availability of a human gene index is assessed as a diseasecandidate gene discovery resource. A feasible approach to construction of a whole-body index required the ability to process error-prone EST data in excess of one million sequences (1,198,607 ESTs as of December 1998). In the absence of new clustering algorithms, at that time, we successfully ported D2_CLUSTER, an EST clustering algorithm, to the high performance shared multiprocessor machine, Origin2000. Improvements to the parallelised version of D2_CLUSTER included: (i) ability to cluster sequences on as many as 126 processors. For example, 462000 ESTs were clustered in 31 hours on 126 R10000 MHz processors, Origin2000. (ii) enhanced memory management that allowed for clustering of mRNA sequences as long as 83000 base pairs. (iii) ability to have the input sequence data accessible to all processors, allowing rapid access to the sequences. (iv) a restart module that allowed a job to be restarted if it was interrupted. The successful enhancements to the parallelised version of D2_CLUSTER, as listed above, allowed for the processing of EST datasets in excess of 1 million sequences. An hierarchical approach was adopted where 1,198,607 million ESTs from GenBank release 110 (October 1998) were partitioned into &quot / tissue bins&quot / and each tissue bin was processed through a pipeline that included masking for contaminants, clustering, assembly, assembly analysis and consensus generation. A total of 478,707 consensus transcripts were generated for all the tissue categories and these sequences served as the input data for the generation of the wholebody index sequences. The clustering of all tissue-derived consensus transcripts was followed by the collapse of each consensus sequence to its individual ESTs prior to assembly and whole-body index consensus sequence generation. The hierarchical approach demonstrated a consolidation of the input EST data from 1,198607 ESTs to 69,158 multi-sequence clusters and 162,439 singletons (or individual ESTs). Chromosomal locations were added to 25,793 whole-body index sequences through assignment of genetic markers such as radiation hybrid markers and g&eacute / n&eacute / thon markers. The whole-body index sequences were made available to the research community through a sequence-based search engine (http://ziggy.sanbi.ac.za/~alan/researchINDEX.html).</p>

Orientador: Mateus José Rodrigues Paranhos da Costa / Coorientador: Fernando Sebastián Baldi Rey / Coorientador: Lucia Galvão de Albuquerque / Banca: Maria Eugênia Zerlotti Mercadante / Banca: Roberto Carvalheiro / Resumo: Este trabalho foi desenvolvido com o objetivo de estudar a associação genética entre características indicadoras de temperamento e de precocidade sexual de fêmeas bovinas da raça Nelore. Foram utilizados dados de temperamento de 7.500 bovinos machos e fêmeas com 18 meses de idade (sobreano) das safras de 2008 e 2009 da Agropecuária Jacarezinho Ltda. (AJ). As características indicadoras de precocidade sexual das fêmeas foram obtidas a partir do arquivo zootécnico da fazenda, para animais nascidos entre os anos de 1984 e 2009. O temperamento foi avaliado durante o manejo de pesagem ao sobreano por meio do teste de movimentação na balança (MOV), atribuindo-se escores de 1 (nenhum movimento) a 5 (animal salta, elevando os membros superiores pelo menos 2,5 centímetros do solo), teste de velocidade de fuga (VF), com uso de um dispositivo de células fotoelétricas que registra o tempo (em segundos) para percorrer uma distância determinada ao sair do tronco de contenção após o manejo de pesagem e o escore de temperamento (ET) realizado pela AJ, atribuindo-se escores 1 (animal calmo) a 5 (animal muito reativo - comportamento agressivo ao observador). Como características indicadoras de precocidade sexual foram utilizadas: a idade ao primeiro parto, em dias (IPP) e a ocorrência de prenhez precoce (PRECO), característica binária, com escore 2 para as novilhas que pariram até 30 meses de idade e escore 1 para as que falharam. Para a estimação dos componentes de (co)variância e parâmetros genéticos para as características estudadas foi utilizada a Inferência Bayesiana. Para VF e IPP foi utilizado um modelo linear e para MOV, ET e PRECO um modelo não-linear (threshold). As estimativas de herdabilidade a posteriori para VF, MOV, ET, IPP e PRECO foram 0,27, 0,11, 0,16, 0,09 e 0,44, respectivamente. As correlações... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The aim of this study was to estimate genetic associations between temperament and female sexual precocity traits in Nellore beef cattle. The temperament was assessed for 7,500 male and female cattle, at 18 months of age (yearling) and born in 2008 and 2009, from the herd of Agropecuária Jacarezinho Ltda. (AJ). The female sexual precocity traits were obtained from the files of the farm for animals born between 1990 and 2009. Temperament was evaluated during the handling for weight determination using a movement score (MOV), assigning scores from 1 (no movement) to 5 (animal jumps, raising the forelegs at least 2.5 cm of the soil); the flight speed (VF), using an electronic device that records the speed at which the animals exit the crush (in m/s) and; temperament score (ET), already in use by AJ, assigning a score from 1 (animal calm) to 5 (animal very reactive - aggressive behavior toward the observer). The female sexual precocity traits used were: age at first calving, in days (IPP) and; occurrence of precocious pregnancy (PRECO), a binary trait in scores of 2 for heifers that calved until 30 months of age and, 1 for the heifers that failed. Bayesian Inference was used for estimation of (co)variance components and genetic parameters. For VF and IPP, a linear model was applied and for MOV, PRECO and ET, a threshold model. The heritability estimates for VF, MOV, ET, IPP and PRECO were 0.27, 0.11, 0.16, 0.09 and 0.44, respectively. The genetic correlations of IPP with VF (0.14), MOV (0.13) and ET (0.09) were all low, as well as for PRECO with VF (-0.19), MOV (-0.03) and ET (-0.03). Although low, all correlations were in a favorable direction, indicating that the temperament is positively associated with sexual precocity. These results suggest that to improve the beef cattle temperament and the sexual precocity, these traits... (Complete abstract click electronic access below) / Mestre

Amplification of chr.1 q21-22 is also an aberration frequently detected in HCC. Copy number gains of the GEF-H1 gene ranked the most frequent event from array-CGH. GEF-H1 up-regulation was significant correlated in patients with advanced HCC staging (P = 0.048), presence of micro-vascular invasion (P = 0.049) and shorter overall and disease free survival of patients (P &lt; 0.03). Similar to BOP1, functional examinations of GEF-H1 suggested profound inhibitory effects on cell motility ( P &lt; 0.035) and invasiveness (P &lt; 0.003) in cell lines studied. Upon GEF-H1 depletion, re-expression of epithelial markers (E-cadherin, cytokeratin 18, alpha-catenin and gamma-catenin) and down-regulations of mesenchymal markers (N-cadherin, fibronectin and vimentin) were also readily observed. In addition, reduced active form of GTP-RhoA together with its downstream effectors including cleaved ROCK 1 and phosphorylated MLC2 were also found in GEF-H1 depleted cells. / Array-CGH also defined candidate proto-oncogenes within 2 causal amplified regions in HCC, chr.8q24 and chr.1q21-q22. In resolving affected genes at chr.8q24, distinctive gains of BOP1 was further established in primary HCC tumors, where frequent BOP1 up-regulations in tumors compared to adjacent non-tumoral liver (P &lt; 0.0001) was identified. Increased BOP1 expression correlated with advanced HCC staging (P = 0.004), micro-vascular invasion (P = 0.006) and shorter overall and disease free survival of patients (P &lt; 0.02). siRNA-mediated suppression of BOP1 in HCC cell lines showed significant inhibition on cell invasion (P &lt; 0.003) and migration (P &lt; 0.05), whereas overexpression of BOP1 in immortalized hepatocyte cell line, L02, showed increase cellular invasiveness and cell migratory rate (P &lt; 0.0001). Evident regression of the Epithelial-to-Mesenchymal Transition (EMT) phenotype was readily identified in BOP1 knockdown cells, where re-expressions of epithelial markers (E-cadherin, cytokeratin 18 and gamma-catenin) and down-regulation of mesenchymal markers (fibronectin and vimentin) were found. It was found that BOP1 likely stimulates actin stress fibers assembly through RhoA activation. / Hepatocellular carcinoma (HCC) is a highly malignant tumor that is associated with a high incidence of cancer morbidity and mortality. Elucidation of genomic aberrations of HCC holds much importance in understanding the molecular basis that underlies the disease causation and progression. Extensive research on HCC has by now revealed a number of key genomic aberrations but, for most of these loci, the underlying cancer-related gene(s) remains unknown. / In this thesis, array-based comparative genomic hybridization (array-CGH) was deployed to define target genes within HCC-associated chromosomal regions. The first part of my study focused on mapping the homozygous deletions (HDs) in HCC. Though infrequent, HD screening has been widely utilized to define tumor suppressor genes (TSGs) in cancers. A panel of HCC cell lines was systematically examined for the presence of HDs. Array-CGH identified 6 HD regions, amongst which CRYL1 (located on chr.13q12.11) displayed most common down-regulations in primary HCC tumors. Significant associations could also be drawn between repressed CRYL1 and advanced tumor staging, increased tumor size and shorter disease-free patient survival (P &le; 0.037). Moreover, HD on CRYL1 could be detected in 36% of HCC cases with CRYL1 down-regulations. Examination of other inactivating mechanisms suggested histone deacetylation and promoter hypermethylation to be likely inactivating events as well. Re-expression of CRYL1 in SK-HEP1 cell line induced profound inhibition on cellular proliferation and cell growth (P &le; 0.002). By Annexin V staining, CRYL1 restoration readily increased pro-apoptotic cells with an induction of P ARP cleavage. Flow cytometry further revealed CRYL1 could prolong the G2-M phase, possibly through interrupting the Cdc2/cyclin B path. / The similarities in functional behaviours of BOP1 and GEF-H1 might have implications in the fundamental biology of HCC tumorigenesis. It is known that HCC is a highly aggressive tumor often associated with intra- and extra-hepatic metastasis. The finding of 2 causal changes to be closely associated with cell migration and invasiveness may have implications in the metastatic potentials of HCC cells being predisposed earlier on from genomic events. / Cheng, Kit Chong Ibis. / Adviser: Nathalie Wong. / Source: Dissertation Abstracts International, Volume: 73-06, Section: B, page: . / Thesis (Ph.D.)--Chinese University of Hong Kong, 2011. / Includes bibliographical references (leaves 177-190). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [201-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.

The human brain is the most complex structure in the known universe and one of the ultimate goals of humanity is to understand its function. The "bottom-up" approach to developmental neuroscience seeks to assemble a "parts list" of the genes expressed in each neuron and a map of the gene regulatory networks that determine the identity of the diverse neuronal types. A key part of building such a gene regulatory map is to identify the transcription factors that are key nodes in these networks. The goal of my PhD was to study the particular gene regulatory networks that govern the decision of the V5 skin cell to divide, lose its skin fate and decide to make dopamine and glutamate sensory neurons. We chose an unbiased forward genetic screen approach coupled with whole genome sequencing of mutants derived from these screens. In the process, we found several mutants that govern this process and developed a software pipeline that simplifies the analysis of mutants for others who perform forward genetic screens.

Kwok, Tsz Piu. / Thesis M.Phil. Chinese University of Hong Kong 2015. / Includes bibliographical references (leaves 109-120). / Abstracts also in Chinese. / Title from PDF title page (viewed on 04, January, 2017).

In this dissertation, the posterior probability of linkage (PPL) framework is extended to the analysis of case-control (CC) data and three new linkage disequilibrium (LD) statistics are introduced. These statistics measure the evidence for or against LD, rather than testing the null hypothesis of no LD, and they therefore avoid the need for multiple testing corrections. They are suitable not only for CC designs but also can be used in application to family data, ranging from trios to complex pedigrees, all under the same statistical framework, allowing for the unified analysis of these disparate data structures. They also provide the other core advantages of the PPL framework, including the use of sequential updating to accumulate LD evidence across potentially heterogeneous sets of subsets of data; parameterization in terms of a very general trait likelihood, which simultaneously considers dominant, recessive, and additive models; and a straightforward mechanism for modeling two-locus epistasis. Finally, being implemented within the PPL framework, the new statistics readily allow linkage information obtained from distinct data, to be incorporated into LD analyses in the form of a prior probability distribution. Performance of the proposed LD statistics is examined using simulated data. In addition, the effects of key modeling violations on performance are assessed. These statistics are also applied to a previously published type 1 diabetes (T1D) family dataset with a few candidate genes with previously reported weak associations, and another T1D CC dataset also previously published as a genome-wide association (GWA) study with some strong associations reported. The new LD statistics under the PPLD framework confirm most of the findings in the published work and also find some new SNPs suspected of being associated with T1D. Sequential updating between the family dataset and the CC dataset dramatically increased the association signal strength for a CTLA4 SNP genotyped in both studies. Linkage information gleaned from the family dataset is also combined into the LD analysis of the CC dataset to demonstrate the utility of this unique feature of the PPL framework, and specifically for the new LD statistics.

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[Truncated abstract] Under prolonged wet and damp conditions, barley grain with low dormancy can germinate precociously, a condition known as preharvest sprouting that causes a number of detrimental effects in grain quality. In particular, preharvest sprouting renders the grain unsuitable for malting. The aim of this study was to take a genomics approach to identify and characterise candidate genes that could be linked to the control of seed dormancy in barley. This thesis developed a bioinformatic strategy that exploited the availability of gene sequences with functional evidence in the model species of Arabidopsis and rice. The bioinformatic strategy integrated phenotypic data (QTL data) and comparative genomics for a targeted approach in identifying candidate genes with a high probability of having a conserved function in cereals. This bioinformatic study identified two candidate genes ERA1 and ABI2 with strong evidence for a role in seed dormancy based on their function in Arabidopsis in abscisic acid (ABA) signal transduction and their co-location to seed dormancy QTLs in Arabidopsis, rice and wheat. In order to establish whether the candidate genes mapped to seed dormancy QTLs in barley, QTL analyses were performed on a double haploid population, not previously studied, developed from a cross between Stirling, a major Australian malting cultivar, and Harrington, a major Canadian malting cultivar. This cross was specifically chosen for this study, as elucidation of chromosomal regions associated with seed dormancy in the background of a malting cultivar would make a significant contribution for the malting industry. '...' Identification of a seed dormancy QTL on the long arm of 3H, in a region syntenic to the wheat chromosome locations of ESTS aligning to the ERA1 and ABI2 genes, laid the foundation for physical and genetic mapping of the candidate genes to investigate whether the genes co-located to the QTL on 3H. Physical mapping of the genes in wheat barley addition lines confirmed their positions on the long arm of 3H. Genetic mapping of the ERA1 gene was performed using a CAPS marker developed in this thesis. The genetic mapping of the ERA1 gene did not place the gene within either of the minor QTLs on 3HL, although segregation distortion may have influenced the map position of this gene. Further investigation is required to resolve the positioning of the ERA1 and ABI2 genes in relation to the 3H seed dormancy QTL. The main outcomes of this study have been 1) identification of candidate genes for further study; 2) identification of QTLs on the long arm of 3H that were previously unknown; 3) demonstration of the potential differences in dormancy that can be achieved through the use of specific gene combinations, highlighting the importance of minor genes and the epistatic interactions that occur between them and; 4) the development of a CAPS marker for the ERA1 gene, which can be used to track the gene in barley breeding programs to observe its association with important agronomic traits. This thesis also pioneered the implementation of several new technologies including multiplex-ready PCR (Hayden et al. 2008) for fluorescence–based SSR genotyping and QTLNetwork (Yang et al. 2008) for statistical analysis of QTLs. Seed dormancy is a complex trait and is likely to involve the interplay of a number of genes that have a role in other developmental and regulatory processes.