Le neuroblastome (NB) est la tumeur solide extracranienne la plus fréquente et mortelle chez les jeunes enfants. Il se caractérise par une résistance à la chimiothérapie possiblement en partie dû à la présence de cellules initiatrices de tumeurs (TICs). Des études ont mis en évidence le rôle de CD133 comme un marqueur des TICs dans divers types de cancers. Les buts de notre travail étaient d’abord de démontrer les vertus de TICs des cellules exprimant CD133 et ensuite, en utilisant une analyse globale du génome avec des polymorphismes nucléotidiques simples (SNPs), d’effectuer une analyse différentielle entre les TICs et les autres cellules du NB afin d’en identifier les anomalies génétiques spécifiques. Des lignées cellulaires de NB ont été triées par cytométrie de flux afin d’obtenir deux populations: une enrichie en CD133 (CD133high), l’autre faible en CD133 (CD133low). Afin de déterminer si ces populations cellulaires présentent des propriétés de TICs, des essais sur les neurosphères, les colonies en agar mou et les injections orthotopiques de 500 cellules sélectionnées dans 11 souris ont été réalisées. Après une isolation de l’ADN des populations sélectionnées, nous avons effectué une analyse génotypique par SNP utilisant les puces « Affymetrix Genome-Wide Human SNP Array 6.0 ». Pour vérifier l’expression des gènes identifiés, des Western Blots ont été réalisés. Nos résultats ont démontré que la population CD133 avait des propriétés de TICs in vitro et in vivo. L’analyse génotypique différentielle a permis d’identifier deux régions communes (16p13.3 and 19p13.3) dans la population CD133high ayant des gains et deux autres régions (16q12.1 and 21q21.3) dans la population CD133low possédant des pertes d’hétérozygoties (LOH). Aucune perte n’a été observée. Parmi les gènes étudiés, l’expression protéique d’éphrine-A2 était corrélée à celle de CD133 dans 6 tumeurs et 2 lignées cellulaires de NB. De plus, l’augmentation de la concentration d’anticorps anti-éphrine-A2 dans le milieu diminue la taille des neurosphères. Ainsi, la population CD133high, qui a des vertus de TICs, possède des caractéristiques génotypiques différentes par rapport à celle CD133low. La présence d’éphrine-A2 dans les cellules exprimant CD133 souligne son importance dans le développement des TICs. Ces résultats suggèrent la présence de potentielle cible pour de nouvelles thérapeutiques ciblant les TICs mise en évidence par l’étude génomique. / Neuroblastoma (NB) is the most common and deadly extracranial solid tumor of childhood characterized by a resistance to chemotherapy possibly due to the presence of tumor initiating cells (TICs). Studies showed the role of CD133 as a marker of TICs in various types of cancers. Our goals were first to demonstrate the stemness of TICs expressing CD133 and then, using a global genomic analysis with single nucleotide polymorphism (SNPs), to perform a differential analysis between TICs and other cells of NB to identify the specific genetic abnormalities. NB cell lines were sorted by flow cytometry to obtain two populations: one enriched in CD133 (CD133high), the other low in CD133 (CD133low). To determine whether these cell populations have TICs properties, we test the ability of cells to form either neurosphères or, colonies in soft agar and we also test their carcinogenic properties by orthotopic injections of 500 selected cells in 11 mice. After a DNA extraction on selected populations, a differential genotyping analysis has been made with Affymetrix Genome-Wide Human SNP Array 6.0. To verify the expression of the genes identified, Western blots had been made. Our results have demonstrated that CD133high population presented TICs properties in vitro and in vivo. The differential genotyping analysis allowed identifying two gains common regions (16p13.3 and 19p13.3) in CD133high population and two others loss of heterozygosity (LOH) (16q12.1 and 21q21.3) in CD133low population . No losses were observed. Among the genes studied, ephrin-A2 protein expression was correlated to CD133 expression in 6 NB tumors and 2 NB cell lines. Also, ephrin-A2’s increased concentration influenced the neurospheres by decreasing their size. Thereby, CD133high population, which had TICs properties, possess different genotyping characteristics compared to CD133low population. The presence of ephrine-A2 in cells expressing CD133 emphasizes its importance in the development of TICs. These results suggest the presence of potential target for new therapies targeting the TICs demonstrated by the genomic study.

L'analyse chromosomique sur puce ADN (ACPA) tend à devenir le principal examen diagnostique dans la déficience intellectuelle (DI). Parmi les techniques d'ACPA, les puces SNP ont l'intérêt de pouvoir détecter les pertes d'hétérozygotie, et par conséquent d'identifier les isodisomies uniparentales (iUPD) et les zones d'identité liées à la consanguinité. Nous avons étudié une cohorte de 1 187 patients atteints de DI, dans un cadre diagnostique, sur puces SNP. Nous avons réalisé, par cette étude, 145 diagnostics (12%) dont 2 iUPD et 6 délétions n'incluant qu'un seul gène. De plus, nous avons détecté 639 CNV rares non décrits chez des sujets contrôles et incluant des séquences codantes, ce qui nous a permis d'identifier 11 gènes candidats dans la DI : CAMTA1, SP3, CNTNAP4, NUDT12, STXBP6, DOCK8, DOCK10, SMARCA2, NYAP2, ATAD3A et ATAD3B. Nous avons tenté de valider l'implication de ces gènes par séquençage, mais n'avons trouvé de seconde mutation pour aucun d'entre eux. Toutefois, des réarrangements de CAMTA1 ont été retrouvés dans 2 autres familles avec un phénotype homogène (DI et ataxie congénitale) ce qui nous a permis d'affirmer qu'il s'agit d'un gène de DI. Par ailleurs, l'homozygosity mapping, réalisé avec puces SNP, a identifié, par séquençage whole exome, une mutation non-sens homozygote du gène BUD13 dans une famille de DI syndromique. Enfin, de façon fortuite, nous avons caractérisé en ACPA une translocation familiale entraînant une disruption d'un gène d'ataxie spino-cérébelleuse, ATXN10, ce qui a permis de mieux comprendre la physiopathologie de cette maladie. Au total, notre étude démontre l'intérêt des puces SNP dans la DI, d'une part en diagnostic et d'autre part pour l'identification de nouveaux gènes responsables de DI.

La vigne est soumise à de nombreuses maladies impliquant l'utilisation de produits phytosanitaires en grande quantité dont l'utilisation est néfaste pour l'environnement et la santé des utilisateurs. Un enjeu est donc de développer des méthodes alternatives à la lutte chimique. La protéine codée par le gène NPR1 (Nonexpressor of pathogenesis-related gene 1) joue un rôle clef dans la résistance à large spectre chez les plantes. Des éliciteurs tels que l'acide salicylique ou des agents pathogènes influencent l'activation de NPR1 dans le cytoplasme. La translocation de NPRl dans le noyau et son interaction avec des facteurs de transcription TGA induit l'expression des gênes PR (Pathogenesis-related). Nous avons identifié sept homologues potentiels des gènes NPR1 et TGA chez Vitis vinifera (VvNPR1.1, VvNPR1.2, VvTGA1 à 5). L'étude de la diversité de séquences dans les exons de 15 accessions de Vitaceae indique qu'ils sont soumis à une forte pression de sélection purificatrice. De plus, l'analyse in silico des régions promotrices des VvNPR1 montre la présence, d'éléments cis-régulateurs potentiels, en réponse aux stress biotiques et abiotiques ainsi que des motifs de liaison à des facteurs de transcription. Une étude plus poussée des introns montre quelques éléments transposables et un faible polymorphisme dans six accessions de Vitis vinifera. Ces résultats argumentent en faveur d'une pression de sélection forte agissant sur ces gènes. Ceci nous a mené à formuler des hypothèses fonctionnelles et à réaliser une étude d'interaction avec les facteurs de transcription VvTGA1 et VvTGA4 par la technique du double hybride. Ces derniers n'interagissent pas avec VvNPR 1.1.

This dissertation should supplement the knowledge of the genetic coronary heart disease (CHD) architecture by analysing known common variation as well as finding novel associated loci and genes. Genotype and allele frequencies, also haplotype blocks were determined in the Lithuanian population according to the list of 60 SNPs associated with CHD. The number of risk alleles per person was determined. The intrapopulation comparison of allele frequencies and haplotype blocks was performed. Allele frequencies were compared with different populations of European ancestry. The association analyses for new loci and candidate gene identification were performed. It was revealed that the common and unique genetic disease architecture variants for different populations exist. Regarding the genetic structure and diversity of the risk SNPs of CHD, Lithuanian population as compared with the European populations falls into the Northern-Southern gradient showing that the Lithuanian gene pool could have experienced the geographical climate and ecological influence during the evolution process. Considering the risk allele number per person of the investigated SNPs, the majority of Lithuanian population individuals have a relatively average risk of developing CHD. Association analyses showed eight new loci associated with CHD, four of which were confirmed and ITPR2 and FBXL17 were found to be the putative candidate genes that could participate in the pathogenesis of CHD and atherosclerosis. / Siekiant papildyti žinias apie genetinę koronarinės širdies ligos (KŠL) architektūrą, užsibrėžta atlikti žinomų genetinių sričių siejamų su KŠL analizę ir naujų genetinių sričių siejamų su šia liga paiešką Lietuvos populiacijoje. Pagal atrinktas žinomas 60 genetinių sričių, siejamų su rizika susirgti KŠL, tiriamiems asmenims nustatyti genotipų ir alelių dažniai, sukonstruoti haplotipų blokai, atliktas vidupopuliacinis alelių dažnių ir haplotipų blokų palyginimas, tarppopuliacinis alelių dažnių palyginimas, nustatytas rizikos alelių tenkančių asmeniui pasiskirstymas populiacijoje. Nustatyta, kad skirtingoms populiacijoms yra būdingi tiek bendri, tiek ir unikalūs genetinės architektūros vienetai, o bendros Lietuvos lietuvių populiacijos genetinę struktūrą ir įvairovę pagal tirtus žymenis galima orientuoti gradientiškai Europos Šiaurės-Pietų kryptimi, tai aiškinant geografinių platumų lemiama klimato ir ekologijos įtaka genofondui evoliucijos eigoje. Pagal tirtų rizikos alelių skaičių tenkantį asmeniui, daugiausia asmenų tirtoje Lietuvos lietuvių populiacijoje turi vidutinę riziką susirgti KŠL. Genetinės asociacijos analize nustatytos aštuonios potencialios naujos kandidatinės su KŠL asocijuotos genetinės sritys, iš jų patvirtintos – keturios, o ITPR2 ir FBXL17 genai yra galimi genai kandidatai dalyvaujantys KŠL ir aterosklerozės patogenezėje.

Siekiant papildyti žinias apie genetinę koronarinės širdies ligos (KŠL) architektūrą, užsibrėžta atlikti žinomų genetinių sričių siejamų su KŠL analizę ir naujų genetinių sričių siejamų su šia liga paiešką Lietuvos populiacijoje. Pagal atrinktas žinomas 60 genetinių sričių, siejamų su rizika susirgti KŠL, tiriamiems asmenims nustatyti genotipų ir alelių dažniai, sukonstruoti haplotipų blokai, atliktas vidupopuliacinis alelių dažnių ir haplotipų blokų palyginimas, tarppopuliacinis alelių dažnių palyginimas, nustatytas rizikos alelių tenkančių asmeniui pasiskirstymas populiacijoje. Nustatyta, kad skirtingoms populiacijoms yra būdingi tiek bendri, tiek ir unikalūs genetinės architektūros vienetai, o bendros Lietuvos lietuvių populiacijos genetinę struktūrą ir įvairovę pagal tirtus žymenis galima orientuoti gradientiškai Europos Šiaurės-Pietų kryptimi, tai aiškinant geografinių platumų lemiama klimato ir ekologijos įtaka genofondui evoliucijos eigoje. Pagal tirtų rizikos alelių skaičių tenkantį asmeniui, daugiausia asmenų tirtoje Lietuvos lietuvių populiacijoje turi vidutinę riziką susirgti KŠL. Genetinės asociacijos analize nustatytos aštuonios potencialios naujos kandidatinės su KŠL asocijuotos genetinės sritys, iš jų patvirtintos – keturios, o ITPR2 ir FBXL17 genai yra galimi genai kandidatai dalyvaujantys KŠL ir aterosklerozės patogenezėje. / This dissertation should supplement the knowledge of the genetic coronary heart disease (CHD) architecture by analysing known common variation as well as finding novel associated loci and genes. Genotype and allele frequencies, also haplotype blocks were determined in the Lithuanian population according to the list of 60 SNPs associated with CHD. The number of risk alleles per person was determined. The intrapopulation comparison of allele frequencies and haplotype blocks was performed. Allele frequencies were compared with different populations of European ancestry. The association analyses for new loci and candidate gene identification were performed. It was revealed that the common and unique genetic disease architecture variants for different populations exist. Regarding the genetic structure and diversity of the risk SNPs of CHD, Lithuanian population as compared with the European populations falls into the Northern-Southern gradient showing that the Lithuanian gene pool could have experienced the geographical climate and ecological influence during the evolution process. Considering the risk allele number per person of the investigated SNPs, the majority of Lithuanian population individuals have a relatively average risk of developing CHD. Association analyses showed eight new loci associated with CHD, four of which were confirmed and ITPR2 and FBXL17 were found to be the putative candidate genes that could participate in the pathogenesis of CHD and atherosclerosis.

The studies in this thesis describe the application of genotyping and allele specific expression analysis to genetic studies. The role of the gene NPC1 in Triglyceride metabolism was explored in mouse models and in humans on the population level in study I. NPC1 was found to affect hepatic triglyceride metabolism, and to be relevant for controlling serum triglyceride levels in mice and potentially in humans. In study II the utility of the HapMap CEU samples was investigated for tagSNP selection in six European populations. The HapMap CEU was found to be representative for tagSNP selection in all populations while allele frequencies differed significantly in the sample from Kuusamo, Finland. In study III the power of Allele specific expression as a tool for the mapping of cis-regulatory variation was compared to standard eQTL analysis, ASE was found to be the more powerful type of analysis for a similar sample size. Finally ASE mapping was applied to regions reported to harbour long non-coding RNAs and associated SNPs were compared to published trait-associations. This revealed strong cis-regulatory SNPs of long non-coding RNAs with reported trait or disease associations.

The possibility of identifying single nucleotide polymorphisms (SNPs) that would be useful for rapid bacterial typing was investigated. Neisseria meningitidis was the organism chosen for modelling the approach since informative SNPs could be found amongst the sequence data available for multi-locus sequence typing (MLST) at http://www.mlst.net. The hypothesis tested was that a small number of SNPs located within the seven gene fragments sequenced for MLST provide information equivalent to MLST. Preliminary investigations revealed that a small number of SNPs could be utilised to highly discriminate sequence types (STs) of clinical interest. Laboratory procedures demonstrated that SNP fingerprinting of N. meningitidis isolates is achievable. Further tests showed that laboratory identification of a defining SNP in the genome of isolates was to be a practical method of obtaining relevant typing information. Identification of the most discriminating SNPs amongst the ever-increasing amount of MLST sequence data summoned the need for computer-based assistance. Two methods of SNP selection devised by the author of this thesis were translated into computer-based algorithms by contributing team members. Software for two computer programs was produced. The algorithms facilitate the optimal selection of SNPs useful for (1) distinguishing specific STs and (2) differentiating non-specific STs. Current input information can be obtained from the MLST database and consequently the programs can be applied to any bacterial species for which MLST data have been entered. The two algorithms for the selection of SNPs were designed to serve contrasting purposes. The first of these was to determine the ST identity of isolates from an outbreak of disease. In this case, isolates would be tested for their membership to any of the STs known to be associated with disease. It was shown that one SNP per ST could distinguish each of four hyperinvasive STs of N. meningitidis from between 92.5% and 97.5% of all other STs. With two SNPs per ST, between 96.7% and 99.0% discrimination is achieved. The SNPs were selected from MLST loci with the assistance of the first algorithm which scores SNPs according to the number of base mismatches in a sequence alignment between an allele of an ST of interest and alleles belonging to all other STs at a specified locus. The second purpose was to determine whether or not isolates from different sources belong to the same ST, regardless of their actual ST identity. It was shown that with seven SNPs, four sample STs of N. meningitidis could, on average, be discriminated from 97.1% of all other STs. The SNPs were selected with the aid of the second algorithm which scores SNPs at MLST loci for the relative frequency of each nucleotide base in a sequence alignment as a measure of the extent of their polymorphism. A third algorithm for selecting SNPs has been discussed. By altering the method of scoring SNPs, it is possible to overcome the limitations inherent in the two algorithms that were utilised for finding SNPs. In addition, the third approach caters for finding SNPs that distinguish members of a complex from non-members.

Aquaculture plays an important role in economic development and food security in many countries in the world. World aquaculture production in 2006 was 51.7 million tonnes with an estimated value of US$ 78.8 billion (FAO, 2009). World production will need to increase however by 30-40 million tonnes from its current production level by 2030 to meet growing global demand for fish. In this context, aquaculture in Vietnam has developed rapidly over the past decade and the fisheries sector ranked fourth in terms of export value in 2008 (Vietnamnet, 2008). Total fisheries production in Vietnam in 2007 was 4.149 million tonnes, of which fisheries production from catch and aquaculture were 2.064 and 2.085 million tonnes, respectively. A variety of aquatic species are cultured in Vietnam, but shrimps (mainly Black Tiger shrimp Penaeus monodon, and Pacific white shrimp Litopenaeus vannamei) and ‘tra’ or ‘basa’ catfish are the most common species used in aquaculture. The giant freshwater prawn (GFP), Macrobrachium rosenbergii, is one of the most important crustacean species in inland aquaculture in many countries across the world where this species is either native or exotic. GFP is suitable for culture in a variety of farming systems, including monoculture or polyculture in ponds, pens, and integrated or rotational rice-prawn culture models. The GFP industry worldwide relies totally on wild or unimproved stocks, a practice that threatens the long-term sustainability of GFP farming due to low productivity and vulnerability of farmed stocks to disease. The current status of GFP aquaculture highlights the need for initiation of a systematic stock improvement program for the species to improve economically important traits. Large-scale selective breeding programs have been instigated for some finfish, salmonids and GIFT tilapia for example, and some selective breeding trials have been conducted on crustacean species, namely marine penaeid shrimp and freshwater crayfish. Examples of selective breeding programs on aquatic species have demonstrated that significant genetic gains can be achieved for growth rates with gains of around 10-20% per generation. While a selective breeding program is an option for GFP stock improvement, an alternative approach to improving GFP productivity, potentially with more immediate effect and one that is less expensive, is crossbreeding which may produce heterosis or hybrid vigour in crossbred offspring. Therefore, a crossbreeding strategy was trialed in the current study as a starting point for a stock improvement program for the GFP industry in Vietnam. The current study assessed the growth performance of three GFP strains (two wild Vietnamese strains from the Dong Nai and Mekong rivers, and a single domesticated Hawaiian strain) and their reciprocal crosses in a complete 3x3 diallel cross, i.e. three purebred and six crossbred strains. The diallel cross was carried out over two consecutive generations (G1 and G2). Juveniles for the experiments were produced using single-pair matings. Juveniles from each strain combination were stocked into three replicate hapas for 15 weeks. Growth data (body weight, carapace length, standard length) from the G1 and G2 were pooled for all subsequent analyses as there was no effect of generation on growth traits. Results showed that the Hawaiian strain performed best among purebred strains, and crosses with the Dong Nai or Mekong strains as dams and the Hawaiian strain as sires grew significantly faster than did the purebred Dong Nai or Mekong strains. These results suggest potential for heterosis among some crosses. Growth data were analyzed in depth by partitioning the strain combination (cross) effect into three components: strain additive genetic effects, heterotic effects, and strain reciprocal effects. Strain additive genetic and reciprocal effects were significant sources of variation for all growth traits measured. Strain additive genetic effects were highest for the Hawaiian strain and lowest for the Mekong strain for all growth traits. Reciprocal effects influenced negatively on growth rate of crosses with the Hawaiian (H) strain as dams and the Dong Nai (D) or Mekong (M) as sires compared with their reciprocal crosses (DH and MH). Heterotic effects for all growth traits were small and not significantly different from zero (P > 0.05). These results indicate that a crossbreeding approach based on the strains evaluated here provides only limited potential for improving growth rates based simply on heterotic outcomes and that a likely more productive option would be to trial artificial selection on a diverse synthetic stock. The current study also employed genetic markers (microsatellites) to characterize levels and patterns of genetic diversity in three purebred strains of GFP that originated from the diallel cross above. All three purebred strains showed relative high levels of genetic diversity in terms of allele number and individual heterozygosity across the six marker loci screened. Levels of genetic diversity present in the three purebred strains combined into a single stock were compared with that from a combination of three wild river stocks to assess the impact of domestication on genetic diversity of a ‘synthetic’ population. Results demonstrated that there was no significant loss of genetic diversity in the three purebred strains combined compared with a reference set containing the three wild populations. Therefore, a synthetic population formed from these purebred strains successfully captured the majority of genetic variation present in the wild broodstock. This synthetic population provides a potential stock for a future selective breeding program for GFP in Vietnam. The current study was also the first attempt to identify single nucleotide polymorphisms (SNPs) in key growth genes in GFP. Two key candidate genes were targeted, actin and crustacean hyperglycemic hormone (CHH), that are potentially linked to growth performance in GFP. The study screened SNPs in GFP females only, because growth performance of GFP males is influenced strongly by social rank. The study identified four SNPs in intron 3 of the CHH gene that were significantly correlated with individual body weight at harvest, while no SNPs detected in the actin gene were associated with growth traits in GFP. This finding however, needs to be confirmed using larger sample sizes and other GFP lines. The current study has produced important basic knowledge relevant to implementation of an effective stock improvement program for GFP in Vietnam. Results indicate that a selective breeding strategy rather than a crossbreeding approach is likely to be the best strategy for improving GFP culture stocks in Vietnam. In addition, the study demonstrates that application of modern molecular genetic technologies can be efficient in developing a genetically diverse, synthetic population for stock improvement and for identifying potential markers correlated with important commercial traits in GFP. Integration of DNA techniques with traditional breeding practices can facilitate GFP stock improvement in Vietnam and accelerate the industry development when improved lines are available. Some limitations of the current study and recommendations for further work are discussed.

Research Doctorate - Doctor of Philosophy (PhD) / Endometrial cancer is one of the most common female cancers in industrialized countries. Traditional risk factors associated with endometrial cancer are well understood and include excessive exposure to estrogen or estrogen unopposed by progesterone. However, variations in the genes that influence these hormones and their association with endometrial cancer have not been well investigated. By studying genetic variation in endometrial cancer, novel markers of risk may be discovered that can be used to identify women at high risk and for the implementation of specialised treatments. Polymorphisms in the genes involved in the following pathways; hormone biosynthesis, hormone receptors, estrogen metabolism, DNA repair and cell cycle control, have been suggested to be involved in the initiation and development of endometrial cancer. The focus of this study was to examine genetic variants in these pathways to assess the existence of an association with the risk of endometrial cancer. In the first part of this study, the COMT V158M polymorphism was examined in a hereditary non-polyposis colorectal cancer (HNPCC) cohort to determine its association with disease expression. The heterozygous genotype was over-represented in women with endometrial/ovarian cancer that did not harbour mismatch repair (MMR) gene mutations. This result suggested that the COMT V158M polymorphism may alter the risk of developing HNPCC related endometrial/ovarian cancer in MMR mutation negative women. Since COMT is involved in the metabolism of estrogen and that estrogen is the main risk factor for endometrial cancer development, closer examination was warranted to determine the association of genetic variation involved in hormone-related pathways and endometrial cancer risk, outside of the context of an inherited predisposition to disease. In the second part of this study, a cohort of 191 women with endometrial cancer and 291 healthy control women were genotyped for polymorphisms in genes involved in hormone biosynthesis, hormone receptors, estrogen metabolism, DNA repair and cell cycle control. The results revealed that variations in estrogen receptor alpha (ESR1) and beta (ESR2), and the androgen receptor (AR), were associated with an increase and decrease in endometrial cancer risk, respectively. Additionally, polymorphisms in CYP1A1, CYP1B1, GSTM1 and GSTP1 were related to a decrease in endometrial cancer risk. A trend was observed for the cyclin D1 870 G>A polymorphism and an increase in endometrial cancer risk, however, this result did not reach significance. Taken together, these results revealed that perturbations in the hormone receptors and estrogen metabolism genes, may aid in the identification of women at high risk of developing endometrial cancer. Interestingly, stratification of the women with endometrial cancer revealed that combinations of polymorphisms in TP53 and MDM2 were associated with higher grades of cancer. This finding may possibly have significant implications as women with reduced apoptotic ability, due to combinations of polymorphisms in these genes, have an increased risk of presenting with higher grades of endometrial cancer, that are associated with lower survival rates. In summary, the results of this thesis showed that variation in the estrogen and androgen receptors, and estrogen metabolism genes, may alter the risk of developing endometrial cancer. Moreover, polymorphisms in the cell cycle control genes, TP53 and MDM2, appear to be associated with higher grades of endometrial cancer. This study of polymorphisms may help explain genetic differences in individual susceptibility to endometrial cancer and are markers of risk that aid in the development of effective and personalised strategies to prevent disease development. This study has improved the understanding of genetic variation associated with endometrial cancer risk. It has the potential to enhance our ability to treat women with endometrial cancer through improved identification and treatment strategies, by virtue of the genetic variation identified, that appears to predispose to disease.

Thesis (MSc)--Stellenbosch University, 2012. / ENGLISH ABSTRACT: Haliotis midae (commonly referred to as perlemoen) is the only one of five endemic species in South Africa that is commercially valued both locally and internationally. Unfortunately, natural perlemoen populations have become a dwindling resource due to commercial exploitation, poaching and the influx of natural threats, such as the West Coast rock lobster, Jasus lalandii. To preserve the natural diversity and sustainability of natural populations as well as commercial stocks, genetic management and improvement of perlemoen is critical. Genetic management requires the utilisation of molecular markers, which aid in the construction of linkage maps and the identification of quantitative trait loci (QTL) associated with economically significant traits. This will allow improvement of commercial stock management in terms of broodstock selection as well as provide valuable insight into natural population dynamics. Single Nucleotide Polymorphisms (SNPs) were selected as the marker of choice due to their successful employment as molecular markers and their wide distribution and abundance within the genomes of various marine species. This study focuses on the characterisation of novel SNPs from transcript sequences generated by Next Generation Sequencing technology. Approximately 40% of the transcripts facilitated the isolation of 105 putative markers, indicating a SNP frequency of ~1% within the H. midae genome. A subset of 24 markers, in addition to 24 previously developed markers, was characterised using the Illumina GoldenGate genotyping assay with the VeraCode technology, a medium to high-throughput genotyping technology. This is the first reported medium- to highthroughput characterisation of SNPs in H. midae. The selected markers were used to determine the efficiency and overall success rate of the GoldenGate platform. Marker characterisation was completed in both natural and commercial populations to determine the utility of these markers for genetic diversity and population structure inference. An 85% genotyping success rate was achieved with the platform. Statistical analysis indicated that the markers developed in this study are suitable for applications including population genetic structure inference, genetic diversity estimation and possibly other downstream applications such as linkage mapping. These markers are considered to be invaluable for future work regarding the genetic management and conservation of H. midae. / AFRIKAANSE OPSOMMING: Haliotis midae (ook bekend as perlemoen) is die enigste van vyf inheemse spesies in Suid-Afrika wat noemenswaardige kommersiële waarde toon plaaslik sowel as internasionaal. Ongelukkig het kommersiële uitbuiting, wildstropery en natuurlike bedreiging (bv. die Weskus kreef Jasus lalandii), wilde perlemoen populasies noemenswaardig verminder. Dus, om natuurlike diversiteit en die voortbestaan van beide wilde en kommersiële populasies te beskerm, is genetiese bestuur en verbetering absoluut noodsaaklik. Genetiese bestuur vereis die gebruik van molekulêre merkers as ’n hulpmiddel in die opstellingvan koppelingskaarte, en die identifisering van die relevante kwantitatiewe eienskap loki (QTL) tipies geassosieer met ekonomies belangrike eienskappe. Die laasgenoemde beoog om kommersiële voorraad bestuur te verbeter, kragtens deur broeidier seleksie sowel as om insig te verskaf m.b.t. wilde bevolking dinamika. Enkel Nukleotied Polimorfismes (SNPs) is gekies as die toepaslike merker vanweë die omvattende toepaslikheid van hierdie merkers binne die genome van verskeie mariene spesies. Hierdie studie fokus op die karakterisering van nuwe SNPs vanuit transkript volgordes ontwikkel deur middel van Volgende Generasie Volgordebepaling (“Next Generation Sequencing”). ’n Beraamde 40% van transkripte het gelei tot die ontwikkeling van 105 potensiëlemerkers, aanduidend van ’n SNP frekwensie van ~1% binne die H. midae genoom. ’n Sub-versameling van 24 merkers, tesame met 24 bestaande merkers, is gekarakteriseer deur die Illumina GoldenGate genotiperings toets met die VeraCode tegnologie, ’n medium tot hoë deurvloei genotiperingstegnologie. Hierdie is die eerste berig van medium tot hoë deurvloei karakterisering van SNPs in H. midae. Die geselekteerde merkers is gebruik om die doeltreffendheid van die GoldenGate platform te bepaal. Merker karakterisering is uitgevoer in beide wilde en kommersiële bevolkings om die effektiewe bruikbaarheid van hierdie merkers m.b.t. genetiese diversiteit, en bevolking struktuur bepaling, te ondersoek. Die platform het ’n 85% genotiperingsukses syfer getoon. Statistiese analise dui daarop dat merkers ontwikkel tydens hierdie studie toepaslik is vir bevolking genetiese struktuur bepaling, genetiese diversiteitberaming en moontlik ook genetiese koppelingskartering. Hierdie merkers word bestempel as onmisbaar vir toekomstige navorsing in genetiese bestuur en bewaring van H. midae.