After the initiation of the human genome sequencing project and the introduction of the field of ‘bioinformatics’, interest in human genetic diversity studies has been increased. Sequence diversity has helped define differences between genes and genomic regions that were previously unknown or difficult to determine. In this thesis I have undertaken to study sequence diversity in the human genome in three areas; 1) investigated diversity in the MHC as represented by the MICA alleles with respect to the known HLA alleles, 2) investigated the structure and diversity in the intergenic region from an MHC related (paralogous) genomic region and related the structural and diversity findings to the knowledge available on the MHC and the wider genome, and 3) described the identification of three and characterization of five new MHC class I polymorphic markers (Alu) and their polymorphic characteristics in worldwide populations and their associations with skin cancer. 1. Phylogenetic analysis of MICA alpha-domain (extracellular) sequences demonstrated relationships with HLA-B cross-reactive serogroups. The HLA-B and MICA loci are in linkage disequilibrium. The data indicated that MICA and HLA-B have evolved in concert from their common ancestors and that the transmembrane polymorphisms have arisen independently and more recently. 2. Sequence analysis of the CD1 genomic region confirmed the presence of five CD1 genes and revealed that there are four unrelated intergenic regions (IGRs). The IGRs are composed mostly of retroelements including five full-length L1 PA sequences and various pseudogenes. Genomic and phylogenetic analyses support the view that the human CD1 gene copies were duplicated prior to the evolution of primates and the bulk of the HLA class I genes found in humans. 3. Five polymorphic Alu insertions (POALINs) were identified (two from previous studies) and located within the 1.8 megabase of the MHC class I genomic region. All five POALINs are polymorphic, and are positively associated with the HLA-A and HLA-B alleles. The AluyHJ insertion was found most frequently associated with HLA-A1 or A24, AluyHG with HLA-A2, AluyHF with HLA-A2, A-10 or -A26 and AluyTF showed a marginal association with HLA-A29. The AluyMICB insertion was strongly associated with HLA-B17 (HLA-B57, HLA-B58) and HLA-B13. The presence of three Alu insertions (AluyHJ, AluyHG and AluyHF) was found in only one HLA class I haplotype (HLA-A1, -B57, -Cw6) in the 10th IHW cell lines. A novel positive association between the presence of AluyMICB and the ‘MICAdel/MICBnull/HLA-B48’ haplotype was determined. The AluyMICB insertion was also associated with at least three different MICB alleles (*0102, *0107N and *0105) and three different HLA-B alleles (B13, B48 and B57). Based on the analysis of associations between different polymorphic markers within the beta block, the MICB*0102 allele was inferred to be the ancestral form of the MICB*0105 and MICB*0107N alleles. The AluyMICB polymorphism can be used to further investigate haplotype relationship and consequently their lineage origins. Some of the MHC POALINs are haplospecific and associate strongly with certain groups of HLA class I alleles and MHC ancestral haplotypes. The AluyTF frequency was significantly associated with skin cancer (p<0.005). MICA gene diversity is derived from two different evolving paths, therefore one or the other alone cannot reliably mark an ancestral haplotype. The CD1 duplicons originated well before the HLA class I duplicons. The MHC POALINs provide new lineage and linkage markers for the fine mapping study of different haplotypes and variations in linkage groups across 1.8 Mb of the MHC class I region. The POALINs may also prove useful in investigating the origins and history of human populations and in determining the role of human genetic diversity in disease risk.

Thesis (Ph. D.)--University of Adelaide, Dept. of Genetics, 1984. / Includes bibliographical references (31 unnumbered leaves at end of vol).

Thesis (Ph.D.)--Murdoch University, 2005. / Thesis submitted to the Division of Science and Engineering. Bibliography: leaves 220-245.

Thesis (M.S. in Biomedical Informatics)--Vanderbilt University, Aug. 2005. / Title from title screen. Includes bibliographical references.

Thesis (Ph. D.) University of Missouri-Columbia, 2006. / The entire dissertation/thesis text is included in the research.pdf file; the official abstract appears in the short.pdf file (which also appears in the research.pdf); a non-technical general description, or public abstract, appears in the public.pdf file. Title from title screen of research.pdf file (viewed on August 3, 2007) Includes bibliographical references.

No description available.

Thesis (PhD)--Stellenbosch University, 2001. / ENGLISH ABSTRACT: Given the organisation of the maize genome as well as demands placed on the saturation of molecular linkage maps it would be desirable to identify informative molecular markers that is located or linked to genic rich areas. Sequences of gene products from different gene classes were investigated. Proteins containing a nucleotide binding site (NBS) and leucine-rich repeat (LRR) region comprise the largest class of disease resistance proteins. Resistant gene analogue (RGA) primers belonging to this specific class were derived from previous published literature studies. By means of similarity studies of short stretches of conserved amino acid and DNA sequences, primers were developed that belonged to the peroxidase and reductase gene classes. A novel class of transposable element was identified, that occurred in the gene rich areas of a diverse range of grass genomes. Of all the MITE families described so far, the Heartbreaker (Hbr) and Hb2 family elements were of particular interest. The unique properties of MITEs, especially their high copy number, polymorphism, stability and preference for genic areas together with the RGA primers, were exploited to develop a new marker technique for the isolation of a class of molecular marker with a strong preference for genic areas. Using the publicly available recombinant inbred population, Tx303 x C0159, 196 MITE/RGA markers were added to the existing recombinant inbred linkage map consisting of ±1033 already established markers. It became apparent that just like loci for disease resistance, the 196 MITE/RGA fragments were not randomly distributed across the maize genome but occurred in clusters spread across the ten maize chromosomes. Ninety-two (92) of the MITE/RGA fragments showed significant correlation to previously mapped maize resistance genes. To establish the conservation and specificity of both the Hbr and Hb2 elements, sequences of 19 MITE/RGA fragments were ascertained. When comparing the partial MITE element sequences from these fragments, a high degree of element conservation was observed. One fragment showed good sequence correlation to a NADPH He Toxin reductase protein product and mapped to the same chromosomal location as the hm1 gene locus in maize. This fragment can be considered a candidate gene for resistance against the pathogen, Helminthosporium carbonum. The Hbr primer used proved to be very specific for the Heartbreaker MITE element, this was in contrast to the non-specificity of the Hb2 primer. The applicability of this technique was tested on two maize diseases that cause immense damage in the maize production industries in South Africa. Fourteen MITE/RGA markers were used to fine map the putative chromosomal locations for the HtN1, Ht1, Ht2 and Ht3 genes that confer resistance. against Setosphaeria turcica, the northern corn leaf blight (NelS) pathogen in maize. Three MITE/RGA fragments were identified that aided in the saturation of the linkage map for quantitative trait resistance (QTl) against gray leaf spot (GlS) in maize. This novel MITE/RGA technique presented a unique opportunity to search for additional candidate genes by using polymerase chain reaction (peR) analysis. When compared to the conventional amplified fragment length polymorphism (AFLP) technique, the MITE/RGA technique proved to be just as efficient but was more cost effective and less time consuming. / AFRIKAANSE OPSOMMING: Die organisasie van die mielie genoom as ook die vereistes wat daar geplaas word op die versadiging van koppelingskaarte, vereis dat daar meer klem geplaas word op die ontwikkeling van molekulêre tegnieke wat merkers in geenryke areas identifiseer. Die volgordes van geenprodukte, wat behoort tot verskillende geenklasse, is deeglik bestudeer. Proteïenprodukte wat bestaan uit 'n nukleotiedbindingsarea (NBA) en 'n leusienryke herhalende (LRH) area is een van die grootste klasse waaronder siekteweerstandsproteïene sorteer. Polimerase kettingreaksie (PKR) inleiers wat behoort tot hierdie spesifieke klas, is verkry vanuit vorige publikasies. Deur kort gekonserveerde aminosuur en DNS volgordes te vergelyk is inleiers ontwikkel wat behoort tot die peroksidase en reduktase gene klasse. 'n Nuwe klas transponeerbare elemente wat voorkom in die geenryke areas van diverse gras genome, is geïdentifiseer. Van al die miniatuur inversie herhalende transponeerbare elemente (MITE) wat al geïdentifiseer is, is die twee elemente, Heartbreaker (Hbr) en Hb2, van groot belang. Unieke eienskappe van die MITEs, veral hul hoë kopie aantal, polimorfiese-indeks, stabiliteit asook voorkeur vir geenryke areas, tesame met die weerstandsgeen analoë (WGA) inleiers, is gebruik om 'n nuwe merker tegniek te ontwikkel. Hierdie nuwe tegniek identifiseer 'n klas merker wat 'n sterk voorkeur het vir geenryke areas. Deur gebruik te maak van die openbare beskikbare rekombinante ingeteelde (RI) populasie, Tx303 x C0159, is 196 MITE/WGA-merkers gekarteer op die bestaande RIL koppelingskaart, wat alreeds bestaan uit ±1033 gevestigde merkers. Net soos die lokusse vir siekteweerstand het dit geblyk dat hierdie 196 merkers in groepe voorkom wat verspreid is oor die tien mielie chromosome. Twee-en-negentig (92) van die 196 gekarteerde MITE/WGA-merkers het betekenisvolle korrelasie gewys met reeds gekarteerde mielie weerstandsgene. Die volgordes van 19 MITE/WGAfragmente is bepaal om sodoende die spesifisiteit en mate van konservering van die Hbr and Hb2 elemente te bereken. 'n Hoë mate van element konservering is waargeneem. Een fragment het In baie goeie volgorde korrelasie gewys met In NADPH HG toksien reduktase proteïen produk en karteer op dieselfde chromosomale posisie as die hm1 geen lokus. Hierdie fragment kan gesien word as In kandidaatgeen vir weerstand teen die mielie patogeen, Helminthosporium carbonum. Die toepasbaarheid van hierdie tegniek is getoets op twee siekte toestande, wat lei tot groot verliese in die mielie industrie, in Suid-Afrika. Veertien van die MITE/WGAmerkers is gebruik om die waarskynlike chromosomale posisies van die HtN1, Ht1, Ht2 en Ht3 gene, wat weerstand bied teen Setosphaeria turcica, die noordelike mielie blaarvlek (NMBV) patogeen, fyner te karteer. Drie MITE/WGA fragmente is geïdentifiseer wat gehelp het in die versadiging van die koppelingskaart vir die kwantitatiewe kenmerk weerstandbiedenheid (KKW) teen grys blaarvlek (GBV) in mielies. Deur gebruik te maak van polimerase kettingreaksie (PKR) analise, verskaf hierdie tegniek die moontlikheid om te soek vir addisionele kandidaatgene. Hierdie tegniek is ook vergelyk met die konvensionele geamplifiseerde fragment lengte polimorfisme (AFLP) tegniek. Daar is gevind dat die nuwe tegniek net so informatief is, maar wel meer koste effektief en tyd besparend.

Thesis (PhD)--Stellenbosch University, 2014. / ENGLISH ABSTRACT: Mycobacterium tuberculosis (M. tuberculosis) is the causative agent of tuberculosis (TB), a disease that affects millions of people world-wide. The species M. tuberculosis consists of a large number of different strains that can be grouped into at least 40 different known strain families. Many of the strains present with different pathogenic characteristics and host adaptations. The F11 LAM strains and Beijing strains currently have a nearly equal representation in the population of Cape Town, making up a total of 45% of all strains in this setting. The Latin-American Mediterranean (LAM) family of M. tuberculosis is proved to be the cause of a large percentage of TB cases worldwide and it is the predominant strain in high-prevalence regions such as the Western Cape and KwaZulu-Natal regions of South Africa, Zambia, Zimbabwe, and South America. This project aimed to investigate the evolution and biological characteristics of the members of the principle genetic group (PGG) 2 Latin-American Mediterranean (LAM) genotype of M. tuberculosis using a combination of whole genomic and proteomic analyses, coupled to mycobacterial molecular epidemiological techniques. The evolution of M. tuberculosis strain families from the Western Cape Province of South Africa proved to be consistent with previous evolutionary scenarios for M. tuberculosis isolated from other parts of the world. This genome-wide SNP-based phylogeny for the evolution of M. tuberculosis offers novel insight into the unique global representation of the M. tuberculosis isolates from the Western Cape, South Africa. The evolutionary scenario presented confirms six LAM sub-lineages, namely IS6110 RFLP families F9, F11, F13, F14, F15, and F26. A subset of sub-lineage defining SNPs was determined for each of the six LAM sub-lineages. The genomic changes in the LAM genotype strains observed through the SNP analysis presented here mostly occur in the genes involved in the cell wall, cell processes, intermediary metabolism and respiration. The same phenomenon was observed when the non-redundant SNPs of the non-LAM isolates were functionally annotated. The functional classification of the regulated proteins in the representative of the LAM RDRio lineage of M. tuberculosis suggests that proteins involved in the lipid metabolism, intermediary metabolism and respiration may be the key to the pathogenic effectiveness of the RDRio LAM lineage. A combination of the LAM SNP analysis and the LAM RDRio/non-RDRio comparison showed that the overall genomic- and proteomic features involved in the cell wall and cell processes of the LAM genotype differ to a large extent from what is seen in the reference strain, M. tuberculosis H37Rv. This genome wide phylogenetic study is the first of its kind in a South African context, and not only presents a robust phylogeny of the M. tuberculosis strain families, and specifically the LAM lineage, but also gives the first ever insight into the protein differences which distinguishes RDRio and non-RDRio M. tuberculosis strains from each other. / AFRIKAANSE OPSOMMING: Mycobacterium tuberculosis (M. tuberculosis) is die mikrobiese agent wat tuberkulose (TB), 'n siekte wat miljoene mense wêreldwyd affekteer, veroorsaak. Die spesie M. tuberculosis bestaan uit 'n groot aantal verskillende stamme wat in ten minste 40 verskillende bekende stam-families gegroepeer word. Baie van die stamme toon verskillende patogeniese eienskappe en gasheer aanpassings. Die F11 LAM stam en Beijing stam het tans 'n byna gelyke verteenwoordiging in die bevolking van Kaapstad, wat 'n totaal opmaak van 45% van stamme wat in hierdie gebied gevind word. Die Latyns-Amerikaanse Meditereense (LAM) familie van M. tuberculosis is bewys om die oorsaak van 'n groot persentasie van TB-gevalle wêreldwyd te wees, en dit is die oorheersende stam in hoë voorkoms streke soos die Wes-Kaap en KwaZulu-Natal streke van Suid-Afrika, Zambië, Zimbabwe en Suid-Amerika. Hierdie projek het ten doel gehad om die evolusie en biologiese eienskappe van die lede van die basiese genetiese groep (BGG) 2 Latyns-Amerikaanse Meditereense (LAM) genotipe van M. tuberculosis te ondersoek deur gebruik te maak van 'n kombinasie van heel genoom en proteoom analise, gekoppel aan mikobakteriële molekulêre epidemiologiese tegnieke. Die evolusie van M. tuberculosis stam families van die Wes-Kaap Provinsie van Suid-Afrika blyk om in ooreenstemming te wees met vorige evolusionêre scenario's vir M. tuberculosis wat in ander dele van die wêreld geïsoleer is. Die genoom-wye enkelnukleotied polimorfisme-gebaseerde filogenetiese hipotese vir die evolusie van M. tuberculosis bied nuwe insig in die unieke wêreldwye verteenwoordiging van die M. tuberculosis isolate van die Wes-Kaap, Suid-Afrika. Die evolusionêre scenario wat hier aangetoon word bevestig ses LAM sub-lyne, naamlik IS6110 RFLP families F9, F11, F13, F14, F15, en F26. 'n Versameling sub-lyn definiërende enkelnukleotied polimorfismes was bepaal vir elk van die ses LAM sub-afstammelinge. Die genomiese veranderinge wat waargeneem is in die LAM-genotipe isolate deur die enkelnukleotied polimorfisme analise wat hier aangebied word, is meestal in die gene wat betrokke is in die selwand, selprosesse, intermediêre metabolisme en respirasie. Dieselfde verskynsel is waargeneem wanneer die nie-oorbodige enkelnukleotied polimorfismes van die nie-LAM isolate funksioneel geannoteer is. Die funksionele klassifikasie van die gereguleerde proteïene in die verteenwoordiger van die LAM RDRio-lyn van M. tuberculosis dui daarop dat die proteïene wat betrokke is in die lipiedmetabolisme, intermediêre metabolisme en respirasie die sleutel tot die patogene doeltreffendheid van die RDRio-LAM-lyn kan wees. 'n Kombinasie van die LAM enkelnukleotied polimorfisme analise en die LAM-RDRio/nie-RDRio vergelyking het getoon dat die totale genomiese- en proteomiese kenmerke wat verwant is aan selwand en selprosesse van die LAM genotipe tot ʼn groot mate verskil van wat gesien word in die verwysing stam, M. tuberculosis H37Rv. Hierdie genoom-wye filogenetiese studie is die eerste van sy soort in 'n Suid-Afrikaanse konteks, en bied nie net ‗n robuuste filogenie van die M. tuberculosis stam families, en spesifiek die LAM genotipe van M. tuberculosis nie, maar gee ook die eerste keer ooit insig in die proteïen verskille wat RDRio en nie-RDRio M. tuberculosis stamme van mekaar onderskei.

published_or_final_version / Biochemistry / Doctoral / Doctor of Philosophy

published_or_final_version / Biological Sciences / Master / Master of Philosophy