Magister Scientiae - MSc (Biochemistry)
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Item The development and application of informatics-based systems for the analysis of the human transcriptome(University of the Western Cape, 2003) Kelso, Janet; Hide, Winston; Faculty of ScienceDespite the fact that the sequence of the human genome is now complete it has become clear that the elucidation of the transcriptome is more complicated than previously expected. There is mounting evidence for unexpected and previously underestimated phenomena such as alternative splicing in the transcriptome. As a result, the identification of novel transcripts arising from the genome continues. Furthermore, as the volume of transcript data grows it is becoming increasingly difficult to integrate expression information which is from different sources, is stored in disparate locations, and is described using differing terminologies. Determining the function of translated transcripts also remains a complex task. Information about the expression profile – the location and timing of transcript expression – provides evidence that can be used in understanding the role of the expressed transcript in the organ or tissue under study, or in developmental pathways or disease phenotype observed. In this dissertation I present novel computational approaches with direct biological applications to two distinct but increasingly important areas of research in gene expression research. The first addresses detection and characterisation of alternatively spliced transcripts. The second is the construction of an hierarchical controlled vocabulary for gene expression data and the annotation of expression libraries with controlled terms from the hierarchies. In the final chapter the biological questions that can be approached, and the discoveries that can be made using these systems are illustrated with a view to demonstrating how the application of informatics can both enable and accelerate biological insight into the human transcriptome.Item Expression and purification of the novel protein domain DWNN(University of the Western Cape, 2002) Lutya, Portia Thandokazi; Pugh, David J.R.; Faculty of ScienceProteins play an important role in cells, as the morphology, function and activities of the cell depend on the proteins they express. The key to understanding how different proteins function lies in an understanding of the molecular structure. The overall aim of this thesis was the determination of the structure of DWNN domains. This thesis described the preparation of samples of human DWNN suitable for structural analysis by nuclear magnetic resonance spectroscopy (NMR), as well as NMR analysis.Item Low detection of exon skipping in mouse genes orthologous to human genes on chromosome 22(University of the Western Cape, 2002) Chern, Tzu-Ming; Hide, Winston; Faculty of ScienceAlternative RNA splicing is one of the leading mechanisms contributing towards transcript and protein diversity. Several alternative splicing surveys have confirmed the frequent occurrence of exon skipping in human genes. However, the occurrence of exon skipping in mouse genes has not yet been extensively examined. Recent improvements in mouse genome sequencing have permitted the current study to explore the occurrence of exon skipping in mouse genes orthologous to human genes on chromosome 22. A low number (5/72 multi-exon genes) of mouse exon-skipped genes were captured through alignments of mouse ESTs to mouse genomic contigs. Exon-skipping events in two mouse exon-skipped genes (GNB1L, SMARCB1) appear to affect biological processes such as electron and protein transport. All mouse, skipped exons were observed to have ubiquitous tissue expression. Comparison of our mouse exon-skipping events to previously detected human exon-skipping events on chromosome 22 by Hide et al.2001, has revealed that mouse and human exon-skipping events were never observed together within an orthologous gene-pair. Although the transcript identity between mouse and human orthologous transcripts were high (greater than 80% sequence identity), the exon order in these gene-pairs may be different between mouse and human orthologous genes. Main factors contributing towards the low detection of mouse exon-skipping events include the lack of mouse transcripts matching to mouse genomic sequences and the under-prediction of mouse exons. These factors resulted in a large number (112/269) of mouse transcripts lacking matches to mouse genomic contigs and nearly half (12/25) of the mouse multi-exon genes, which have matching Ensembl transcript identifiers, have under-predicted exons. The low frequency of mouse exon skipping on chromosome 22 cannot be extrapolated to represent a genome-wide estimate due to the small number of observed mouse exon-skipping events. However, when compared to a higher estimate (52/347) of exon skipping in human genes for chromosome 22 produced under similar conditions by Hide et al.2001, it is possible that our mouse exon-skipping frequency may be lower than the human frequency. Our hypothesis contradicts with a previous study by Brett et al.2002, in which the authors claim that mouse and human alternative splicing is comparable. Our conclusion that the mouse exon-skipping frequency may be lower than the human estimate remains to be tested with a larger mouse multi-exon gene set. However, the mouse exon-skipping frequency may represent the highest estimate that can be obtained given that the current number (87) of mouse genes orthologous to chromosome 22 in Ensembl (v1 30th Jan. 2002) does not deviate significantly from our total number (72) of mouse multi-exon genes. The quality of the current mouse genomic data is higher than the one utilized in this study. The capture of mouse exon-skipping events may increase as the quality and quantity of mouse genomic and transcript sequences improves.Item Optimizing the ion source for polarized protons(University of the Western Cape, 2005) Johnson, Samantha; Celliers, P.; Faculty of ScienceBeams of polarized protons play an important part in the study of the spin dependence of the nuclear force by measuring the analyzing power in nuclear reactions. The source at iThemba LABS produces a beam of polarized protons that is pre-accelerated by an injector cyclotron (SPC2) to a energy of 8 MeV before acceleration by the main separated-sector cyclotron to 200 MeV for physics research. The polarized ion source is one of the two external ion sources of SPC2. Inside the ion source hydrogen molecules are dissociated into atoms in the dissociator and cooled to a temperature of approximately 30 K in the nozzle. The atoms are polarized by a pair of sextupole magnets and the nucleus is polarized by RF transitions between hyperfine levels in hydrogen atoms. The atoms are then ionized by electrons in the ionizer. The source has various sensitive devices, which influence beam intensity and polarization. Nitrogen gas is used to prevent recombination of atoms after dissociation. The amount of nitrogen and the temperature at which it is used plays a very important role in optimizing the beam current. The number of electrons released in the ionizer is influenced by the size and shape of the filament. Optimization of the source will ensure that beams of better quality (a better current and stability) are produced.