Thesis (MSc (Genetics))--University of Stellenbosch, 2005. / Breast cancer is a major cause of morbidity and mortality as it is the most common invasive cancer in women worldwide. The lifetime risk for South African women to develop breast cancer is one in 31. A family history of the disease is a well-established risk factor and germline mutations in the BRCA1 (breast cancer one) and BRCA2 (breast cancer two) tumour suppressor genes markedly increase the risk of developing breast cancer. A few hundred mutations spanning the entire coding sequences of both genes have already been reported. Numerous other breast cancer susceptibility loci have been identified, but results from association studies are discrepant. The checkpoint kinase gene, CHEK2, and specifically the CHEK2*1100delC variant has, however, consistently been implicated as a candidate low-penetrance breast cancer allele. To date, few comprehensive molecular-genetic studies have been completed for the various South African breast cancer populations. The aim of this study was to determine the BRCA1 and BRCA2 mutation spectrum and prevalence in two South African populations, namely Mixed Ancestry and Caucasian. The frequency of the CHEK2*1100delC mutation was also investigated. The patient group comprised 101 unrelated patients (98 women and 3 men), presenting with invasive breast cancer. Patients with a moderate family history of breast cancer (n=48) were screened for the CHEK2*1100delC allele and the coding sequences of the BRCA1 (partly completed in a previous study) and BRCA2 genes. Patients without a family history of the disease (n=53) were only screened for the CHEK2*1100delC allele. Mutation detection was done using combined single-strand conformation polymorphism and heteroduplex analysis (SSCP/HA), followed by DNA sequencing of the identified variants. Due to its size (~5kb), exon 11 of BRCA2 was sequenced directly after amplification, in seven overlapping fragments. Three deleterious BRCA1 mutations, 1623_1627delTTAAA, E881X and 5313delC have previously been identified in three patients from the study population. No additional pathogenic mutations have been detected in this gene during this study. Two deleterious BRCA2 mutations, 6677_6678insTA and 8162delG, were identified in two and three patients respectively. Overall, BRCA1 and BRCA2 mutations have been identified in 17% of the Mixed Ancestry patients and in 15.8% of the Caucasian patients. Together BRCA1 and BRCA2 mutations account for 16.7% of breast cancer in the study population. In addition, a number of silent polymorphisms as well as variants of unknown functional significance, both known and novel, were identified. The E881X variant, which has been reported as an Afrikaner founder mutation (Reeves et al. 2004), was identified in one patient of Mixed Ancestry, but none of the published European founder mutations have been detected in our patient group. This suggests a unique mutation spectrum for South African breast cancer patients. The prevalence of the BRCA2 mutations, 8162delG and 6677_6678insTA, has to be elucidated within a larger study group. Haplotype analysis will reveal whether these patients have a common ancestor. Our findings do not suggest the presence of the CHEK2 variant in South African breast cancer patients, but a larger study population has to be analysed to confirm this. The results of this study are in agreement with those from other populations, indicating that less than 20% of breast cancers that occur in individuals with a moderate-risk for developing breast cancer are due to BRCA1 and BRCA2 mutations. By determining the contribution of BRCA1 and BRCA2 mutations to breast cancer in this group of patients, one can assess the appropriateness of predictive or diagnostic DNA testing in the clinical setting.

Thesis (MPhil)--University of Stellenbosch, 2004. / ENGLISH ABSTRACT: Tilapia is presumed to have a well-defined genetic mechanism of sex determination, but not all sex ratios are compatible with a monofactorial sex determination model. A theory of autosomal gene influence, as well as temperature sex determination (TSD) has been proposed in order to explain large variations in sex ratios. This study assessed the variation in progeny sex ratio in O. mossambicus as a basis for the application of YY male technology in the production of all-male progeny groups. Three populations of O. mossambicus were sampled that are representative of the geographical distribution of the species in Southern Africa. Progeny groups were produced from randomly selected parents and maintained at constant temperature during the labile period of sex differentiation. Variation in sex ratios between different families of the same strain as well as between different strains was calculated. No significant differences were observed in sex ratio between strains, though a significant intra-group variation was identified. This study identified both male and female-biased sex ratios. The data in general conform to a monofactorial sex determination model. Male-biased sex ratio observed in one strain can possibly be ascribed to temperature sex determination (TSD). This strain of O. mossambicus originated from an area with different annual temperature patterns and the possibility of TSD having an adaptive advantage is discussed. This thesis also presents the results of a program to produce monosex male tilapia through the application of the YY male technology in O. mossambicus. Viable XY female and YY male genotypes were produced. XY females sired progenies ranging from 68-94% male ofspring, while YY males sired a mean progeny of 94% male phenotypes. From these results it is concluded that YY male technology provide a viable method for the production of all-male progeny in O. mossambicus. Once available on a commercial scale, the technology can be made more reliable through the application of the appropriate selection methods. / AFRIKAANSE OPSOMMING: Tilapia spesies is oorwegend onderworpe aan ‘n goed gedefinieerde enkelfaktor model van genetiese geslagsbepaling, hoewel afwykings van sodanige geslagsbepalingsmodel soms waargeneem word. Die invloed van outosomale gene, sowel as omgewings temperatuur word voorgehou as verklaring van die waargenome variasies in geslagsverhoudings. Hierdie studie behels die evaluasie van variasie in geslagsdifferensiasie van O. mossambicus as basis vir die implementering van die YY manlike tegnologie vir die produksie van slegs-manlike nageslag groepe. Monsters is bekom van drie populasies van O. mossambicus verteenwoordigend van die geografiese verspreiding van die spesie in Suider Afrika. Nageslag groepe is geproduseer vanaf ewekansig gekose ouerpare en by ‘n konstante temperatuur gehuisves tydens die sensitiewe tydperk van geslagsdifferentiasie. Variasie in geslagsverhouding tussen verskillende families binne dieselfde subpopulasie sowel as binne verskillende populasies is bepaal. Geen betekenisvolle verskille is waargeneem in die geslagsverhouding tussen die verskillende populasies nie terwyl betekenisvolle verskille tussen families binne ‘n populasie waargeneem is. Die data voldoen aan ‘n enkelfaktor genetiese geslagsbepalingsmodel, met die uitsondering van een populasies waar ‘n moontlike omgewingsverwante geslagsbepalingseffek waargeneem is. Die tesis sluit ook in die resultate in van ’n program om YY-tegnologie in O. mossambicus te vestig ten einde manlike nageslag groepe te produseer. Funksionele vroulike XY- en manlike YY genotipes is ontwikkel. Die vroulike XY-genotipes het manlike nageslag groepe opgelewer wat wissel van 68%-94% manlik, terwyl manlike YY-genotipes nageslag groepe gelewer het wat gemiddeld 94% manlik was. Die resultate bevestig die lewensvatbaarheid van YY-tegnologie vir die kommersiële produksie van manlike nageslaggroepe in O. mossambicus.

Thesis (MSc (Genetics))—University of Stellenbosch, 2007. / In South Africa, pre-eclampsia is the second highest cause of maternal deaths. The incidence of this disease in the Western Cape alone is 6.8% and places a large burden of health care facilities. The placenta and implantation thereof is thought to play the most significant role in the onset of this disease. Among the many theories for its aetiology, is the acknowledged two - stage theory. This is based on evidence that pre-eclamptic placentas demonstrate altered remodelling and invasion into the uterine endometrium and myometrium. The sub-optimal endometrium invasion leads to less oxygenation of the placental environment causing transient hypoxia. Consequently, the placenta is thought to release unknown factors into the maternal circulation which then culminates in clinical features associated with pre-eclampsia. Neurokinin B is thought to be one of these placental factors and subsequently binds to the NKB receptor in the maternal system. Endothelium-derived nitric oxide synthase has recently been shown to activate this receptor. The aim of this study was to investigate the role of neurokinin B (TAC3) and the neurokinin B receptor (TACR3) genes in the predisposition of pre-eclampsia and their interaction with eNOS in the South African coloured population together with a matched control cohort.

Thesis (MScAgric)--Stellenbosch University, 2012. / ENGLISH ABSTRACT: The abalone Haliotis midae is the most important aquaculture species in South Africa. The industry is dependent on export to Far Eastern markets in a variety of forms, including live, frozen, canned or dried. The species is considered undomesticated in the sense that the current commercial broodstock has been obtained from natural populations through a process of random collection. Global competition has necessitated the South African industry to introduce a genetic improvement program to increase biological productivity and financial profitability. The objective of this study was to assess the genetic variation and to estimate key parameters in terms of growth and yield related traits, between family groups that form part of the breeding program. The study reports on heritability estimates of growth rate (0.14 ± 0.05), canning yield (0.08 ± 0.03), and drip loss during live export (0.03 ± 0.02). The high genetic correlation (0.94 ± 0.34) between shell length and live weight enables industry to utilise either weight or shell length as a criteria during operational practices such as sorting, grading and harvesting. The correlation of 0.85 ± 0.01 between live weight and canning loss indicates that animals that weigh more have a lower dressing percentage. Based on these low heritability values obtained for yield related traits it is recommended not to include these traits in the selection program at this stage. The findings of the study were however, compromised by the availability of a limited number of family groups, the age differences between families and the effect of different locations on the variance in phenotypes. Further investigation is needed to confirm the credibility of the results. / AFRIKAANSE OPSOMMING: Die perlemoen Haliotis midae is die belangrikste akwakultuurspesie in Suid Afrika. Die industrie is afhanklik van uitvoere na markte in die Verre Ooste. ‘n Verskeidenheid van produkte word uitgevoer, insluitend lewendige, gevriesde, gedroogte en verblikte perlemoen. Die spesie word as ongedomestikeerd beskou aangesien die huidige teeldiere op ‘n lukrake wyse uit natuurlike populasies versamel is. Globale kompetisie het die Suid Afrikaanse industrie genoodsaak om ‘n genetiese verbeteringsprogram in werking te stel om sodoende die biologiese produktiwiteit en finansiële winsgewendheid te verbeter. Die studie poog om genetiese variasie in groei en opbrengsverwante kenmerke tussen familiegroepe wat deel uitmaak van die teelprogram te ondersoek. Oorerflikheid van groeitempo (0.14 ± 0.05), opbrengs na verblikking (0.08 ± 0.03), en vogverlies na lewendige uitvoer (0.03 ± 0.02) is beraam. Die hoë korrelasie (0.94 ± 0.34) tussen gewig en skulplengte stel die industrie in staat om beide massa en skulplengte as kriteria te geruik tydens operasionele praktyke van sortering, groepering en oes. Die korrelasie van 0.85 ± 0.01 tussen gewig en verlies na verblikking dui aan dat swaarder diere ‘n laer uitslagpersentasie het. As gevolg van die lae oorerflikheidswaardes vir opbrengsverwante kenmerke word daar aanbeveel dat hierdie kenmerke nie op hierdie stadium ingesluit word in die seleksieprogram nie. Resultate is egter beïnvloed deur ‘n beperkte aantal familie groepe, ouderdomsverskille tussen families en die effek van verskillende lokaliteite op die variansie in die fenotipes. Verdere ondersoeke is nodig om die geloofwaardigheid van die bevindinge te bevestig.

Thesis (MScAgric)--Stellenbosch University, 2011. / ENGLISH ABSTRACT: See full text for abstract / AFRIKAANSE OPSOMMING: Sien volteks vir opsomming

Thesis (MSc (Genetics. Institute for Plant Biotechnology))--University of Stellenbosch, 2006. / The main goal of this project was to elucidate the apparent role of neutral invertase (NI) in sucrose accumulation in sugarcane. In the first part of the study putative transgenic cell lines (transformed with antisense NI constructs) were characterised to confirm the stable integration and expression of the transgene. Batch suspension cultures were used to initiate replicate cultures of several of these transgenic lines as well as a control, and the metabolism of the cultures during a 14 day growth cycle was examined. The transgenic lines had substantially reduced levels of NI activity. While the activities of the other invertases remained unchanged, the activity of sucrose synthase (SuSy) was significantly higher in the transgenic suspension cultures relative to the control. Throughout the growth cycle, sucrose concentrations in the transgenic lines were consistently higher, and glucose and fructose concentrations lower, than the control. The transgenic cultures also exhibited a decreased growth rate in comparison to the control. Labelling studies confirmed a decrease in the in vivo rate of invertase-mediated sucrose hydrolysis in the transgenic lines, as well as indicating a decline in the partitioning of carbon to respiratory pathways in these cultures. In the second part of the study, which focussed on greenhouse-grown transgenic plants, similar results were reported. NI activity was significantly decreased, and SuSy activity increased in all of the tissues sampled. The sucrose concentration and purity were also higher in the transgenic tissues, while the in vivo sucrose hydrolysis rate was lower. Allocation of carbon to respiration was lower in the transgenic plants, suggesting that a decrease in sucrose breakdown reduces the availability of hexoses for growth and respiration. Overall, the results suggest that NI plays a key role in the control of sucrose metabolism, and that changes in the activity of this enzyme have far-reaching effects on cellular metabolism. The fact that the trends reported in the whole-plant studies parallel those of the suspension cultures confirms that suspension cultures can be used as a model system in metabolic engineering research in sugarcane. Thus the possibility now exists to analyse large numbers of transgenic lines in a quicker time frame and at a reduced cost in comparison to conventional methods.

Thesis (MSc(Genetics))--University of Stellenbosch, 2009. / ENGLISH ABSTRACT: The hypertensive conditions of pregnancy (including pre-eclampsia (PE)) is the leading cause of primary obstetric death in South Africa and affects at least five percent of pregnancies in the Western Cape province. Reduced levels of placental protein 13 (PP13) early in pregnancy are associated with a higher incidence of PE in later gestation. PP13 and annexin II have been co-localised to the brush border membrane of syncytiotrophoblasts, and form a complex that is transported to the maternal circulation. It is speculated that genetic variation in the gene encoding annexin II (ANXA2) could underlie the reduced PP13 levels. The aim of this study was to screen the ANXA2 gene, including the proximal promoter region, in two South African population groups, (Mixed Ancestry and Black) from the Western Cape, to identify whether variants in the ANXA2 gene confer susceptibility to PE. The study cohort comprised of 120 pre-eclamptic maternal, 94 pre-eclamptic fetal and 54 healthy control individuals. Genomic DNA of patient and control individuals was extracted for PCR amplification of ANXA2 and Multiphor SSCP/HD analysis was performed for mutation detection. The conformational variants identified were subjected to automated DNA sequencing and subsequently to RFLP analysis, to confirm the genotypes in the remainder of the cohort. Nine previously identified variants (c.-31 T>C, c.292 G>T; p.Val98Leu, c.975 C>T;p.Gly325Gly, c.-12+75 C>A, c.-11-43 G>A, c.-11-13 A>T, c.48+67 C>T, c.449-17 G>A, c.683-56 G>A) and 16 novel variants (c.-442 C>G, c.-191 G>C, c.-189_-188insGCCGG, c.-135 C>G, c.-92 A>T, c.222 C>T; p.Ala74Ala, c.600 C>T; p.Asp110Asp, c.934 G>A; p.Gly312Ser, c.244-42 G>C, c.244-76 C>G, c.528+38 C>T, c.589-5 C>T, c.682+49 C>T, c.961-30 A>G, c.961-24 C>G, c.*1057 A>G) were identified upon screening the ANXA2 gene. Statistical analysis identified significant association at five loci: SNP c.-92 A>T located within the ANXA2 5‟UTR, exonic SNP c.222 C>T; p.Ala74Ala and three intronic SNPs c.244-76 C>G, c.449-17 G>A and c.589-5 C>T. Three of the five variants (c.-92 A>T, c.244-76 C>G, c.589-5 C>T) were significantly associated with PE (P<0.05) and could contribute to PE susceptibility in these two SA iv populations, whereas the other two variants (c.222 C>T; p.Ala74Ala, c.449-17 G>A) revealed a possible protective effect, suggesting a reduced risk of developing PE. In silico analysis predicted the disruption and creation of several putative transcription factor binding sites by three SNPs in the ANXA2 gene, which could subsequently affect ANXA2 functioning. This study provides evidence for genetic variation in the ANXA2 gene, which warrants functional experimental validation in an attempt to investigate the function of these SNPs in molecular, cellular and physiological processes underlying PE. Identifying an association between variants in the ANXA2 gene and PE could contribute to the development of an additional early biomarker. The early identification of PE would promote the South African health system by providing the appropriate health care support and monitoring of high risk pregnancies, which could ultimately result in improved pregnancy outcome. / AFRIKAANSE OPSOMMING: Die hipertensiewe siektes van swangerskap (insluitende pre-eklampsie (PE)) is die belangrikste direkte oorsaak van moedersterftes in Suid-Afrika en dit kom voor by ongeveer 5% van swangerskape in die Wes-Kaap provinsie. Verlaagde plasentale proteïen 13 (PP13) vlakke tydens vroeë swangerskap word verbind met „n hoër voorkoms van PE in latere swangerskap. PP13 en anneksin II kom albei op die borselgrens membraan van synsitiotrofoblaste voor waar hulle „n kompleks vorm wat na die moederlike sirkulasie vervoer word. Daar word gespekuleer dat die onderliggende oorsaak vir laer PP13 vlakke as gevolg van genetiese variasie in die geen wat anneksin II kodeer (ANXA2) kan wees. Die doel van hierdie studie was om die ANXA2 geen, insluitende die proksimale promoter area, in twee Suid-Afrikaanse populasie groepe, (Kleurling en Swart) van die Wes-Kaap, te skandeer met die doel om variante in die ANXA2 geen te identifiseer en ‟n moontlike assosiasie met die vatbaarheid vir PE te bepaal. Hierdie studie populasie het bestaan uit 120 pre-eklamptiese vroue, 94 neonate van pre-eklamptiese ma‟s en 54 gesonde kontrole individue. Genomiese DNS van die pasiënte en kontrole individue is geëkstraeer vir polimerase kettingreaksie amplifikasie van die ANXA2 geen, waarna Multiphor enkelstring konformasie polimorfisme heterodupleks analise uitgevoer is met die doel om DNS variante te identifiseer. Die verskillende konformasies waargeneem is onderwerp aan semi-geoutomatiseerde DNS volgorde bepalingsanalise en gevolglik restriksie fragment lengte polimorfisme analise om genotipes in die res van die studiegroep te bevestig. Vyf-en-twintig variante is geïdentifiseer met die skandering van die ANXA2 geen, waarvan nege voorheen geïdentifiseer is (c.-31 T>C, c.292 G>T; p.Val98Leu, c.975 C>T;p.Gly325Gly, c.-12+75 C>A, c.-11-43 G>A, c.-11-13 A>T, c.48+67 C>T, c.449-17 G>A, c.683-56 G>A) en 16 nuwe variante is (c.-442 C>G, c.-191 G>C, c.-189_-188insGCCGG, c.-135 C>G, c.-92 A>T, c.222 C>T; p.Ala74Ala, c.600 C>T; p.Asp110Asp, c.934 G>A; p.Gly312Ser, c.244-42 G>C, c.244-76 C>G, c.528+38 C>T, c.589-5 C>T, c.682+49 C>T, c.961-30 A>G, c.961-24 C>G, c.*1057 A>G). Statistiese analise het „n statisties beduidende assosiasie met vyf SNPs geïdentifiseer: SNP c.-92 A>T geleë in die ANXA2 5‟UTR, die koderende SNP c.222 C>T; p.Ala74Ala en drie SNPs c.244-76 C>G, c.449-17 G>A and c.589-5 C>T geleë in die nie-koderende areas. Drie van hierdie vyf SNPs (c.-92 A>T, c.244-76 C>G, c.589-5 C>T) het statisties beduidende assosiasie met PE (P<0.05) getoon en kan bedra tot die vatbaarheid vir PE in hierdie twee Suid-Afrikaanse populasies, terwyl die ander twee SNPs (c.222 C>T; p.Ala74Ala, c.449-17 G>A) „n moontlike beskermende effek gedui het, wat „n verlaagde risiko vir die ontwikkeling van PE voorstel. In silico analise het voorspel dat verskeie voorgestelde transkripsiefaktor bindingsetels onderbreek of geskep sal word in die teenwoordigheid van drie SNPs in die ANXA2 geen, wat gevolglik die funksionering van ANXA2 kan affekteer. Hierdie studie verskaf bewyse vir genetiese variasie in die ANXA2 geen, wat verdere funksionele eksperimentele ondersoeke vereis om die funksie van hierdie SNPs in molekulêre, sellulêre en fisiologiese prosesse onderliggend aan PE te bepaal. Die identifisering van ‟n assosiasie tussen variante in die ANXA2 geen en PE kan bydra tot die ontwikkeling van ‟n addisionele vroeë genetiese merker. Die vroeë identifisering van PE kan die Suid-Afrikaanse gesondheidsisteem geweldig baat deurdat die geskikte gesondheidsorg en ondersteuning asook deurgaanse monitering van hoë risiko swangerskappe verskaf sal kan word. Dit kan uiteindelik lei tot ‟n verbeterde uitkoms vir swangerskappe in Suid-Afrika.

Thesis (MSc (Genetics))--University of Stellenbosch, 2007. / The aim of this study was to develop a DNA-based diagnostic system that can be used to distinguish between genotypes in the wheat breeding program at the University of Stellenbosch. Known marker systems were investigated and the chosen marker system would then be implemented to determine its utility in the breeding program. Three marker systems were considered, i.e. microsatellites, Amplified Fragment Length Polymorphisms (AFLPs) and various retrotransposon-based markers. Each system is based on polymerase chain reaction (PCR) amplification from specific primer pairs. The multitude of primer options was narrowed down during a review of published literature regarding wheat molecular markers. Thirty nine microsatellite primer pairs and nine AFLP primer combinations were chosen for the initial genotype evaluation. Four different retrotransposonbased techniques were investigated; namely Inter-Retrotransposon Amplified Polymorphism (IRAP), REtrotransposon-Microsatellite Amplified Polymorphism (REMAP), Sequence- Specific Amplified Polymorphism (SSAP) and, a derivative of these developed in this study, Wis-2 Retrotransposon Amplification. The study started with twenty genotypes which included varieties/breeding lines from five breeding programmes. The genotypes were chosen as representative of the respective breeding populations and were used in the initial testing of the marker systems. Eighteen microsatellites were evaluated using the panel of twenty genotypes. From this, six primer pairs (Xgwm190, Xgwm437, Xgwm539, Xwmc11, Xwmc59 and Xwmc177) were chosen to test the semi-automated DNA sequencer detection system. A single band/peak in each microsatellite profile was used for genotyping. Four of the primer pairs were labelled with different fluorochromes which enabled them to be multiplexed. The differences in amplification products of the six microsatellites meant that all six could be detected in one electrophoresis run. The banding pattern produced by microsatellite Xwmc177 was complex and highly polymorphic and was therefore also analysed in the same way as the AFLP patterns. When analyzed in this manner it proved to be more informative than the combination of six microsatellites (with a single prominent band scored in each). Three AFLP primer combinations could also be multiplexed and visualised together. The three EcoRI selective primers were labelled with different dyes and used with one MseI selective primer. The SSAP system also used fluorescently labelled primers and proved to be the most useful of the retrotransposon-based methods. However, this system produced such a large amount of data that it made analysis too time consuming. Therefore the six microsatellites and three AFLP primer combinations (MseI-CTC and EcoRI-ACA, -AAC, - AGG) were selected for routine genotyping. Due to the numerous highly polymorphic bands produced by the SSAP system it could be very useful to differentiate very closely related genotypes that cannot be distinguished with the markers proposed for routine use. A panel of 119 breeding lines were then used to implement the two chosen marker systems. The results obtained for these markers were used to produce a dendrogram of the lines using the SAS cluster analysis function. The clusters showed that most of the lines could be distinguished from each other. The MseI-CTC and EcoRI-AGG primer combination was the most informative. It produced the largest number of clusters (53) and could therefore discriminate between more of the lines than any other method. The dendrograms and clusters allowed sixteen of the breeding lines to be selected to test the optimal number of seeds to represent an entire population (variety/breeding line) as one seed was not sufficient. It was decided that eight seeds could provide a good representation of the intra-line variability.

Thesis (MScAgric)--Stellenbosch University / ENGLISH ABSTRACT: Triploidy is the genetic state of containing three sets of chromosomes per cell in stead of two as in diploid organisms. The South African abalone (Haliotis midae) is naturally a diploid organism that sexually matures between four to eight years of age. Early sexual maturity is a disadvantage in cultured abalone stock, as the process of gonad development and spawning is energy demanding, causing energy to be diverted away from somatic growth. This same problem has been extensively experienced in diploid bivalve molluscs, where triploidy has since been applied as a means to prevent sexual maturation from occurring, thereby speeding up the growth process and shortening the time to marketing. Because triploidy was effective in bivalves, it was thought that it could contribute to faster growth in abalone as well. A procedure for the induction of triploidy in the abalone, Haliotis midae, was developed by De Beer (2004) and yielded up to 100 percent triploidy in treated abalone larvae. The next step was to compare the growth of the diploids and triploids to establish whether there was indeed a growth advantage on the part of the triploids, in view of commercial application. By using the same techniques as described by De Beer (2004), three groups consisting of triploid and diploid siblings were produced and subscribed to a comparative growth trial. The groups were spawned in three different seasons. The main objective was to establish whether there was in fact a difference in growth between diploid and triploid siblings, and whether seasonal effects were associated with growth advantages for either triploids or diploids. The two growth parameters measured were shell length and body weight. Measurements commenced at eight months of age, when the abalone could be individually tagged and continued up to the age of 24 months. The over-all results provided no convincing evidence of statistically significant faster growth of triploid juveniles compared to that of diploids up to two years of age. Growth differences were detected between seasons, but could not confidently be ascribed to seasonal environmental effects. The regression of shell length to body weight was similar for diploids and triploids. / AFRIKAANSE OPSOMMING: Triploiede organismes bevat drie stelle chromosome per sel in plaas van twee soos dit normaalweg in diploiede diere voorkom. Die Suid Afrikaanse perlemoen (Haliotis midae) is van nature ‘n diploiede organisme wat tussen die ouderdom van vier tot agt jaar seksueel aktief word. Vroeë seksuele aktiwiteit is ongewens in kommersiële akwakultuur aangesien energie spandeer word aan gonade ontwikkeling in plaas van somatiese groei. Dieselfde probleem is vroeër in die oester bedryf ondervind waar dit deur middel van triploiede induksie aangespreek is. Triploiedie veroorsaak steriliteit en kan gebruik word as ’n metode om steriliteit op groot skaal te induseer. Steriliteit sou dan meebring dat meer energie beskikbaar is vir somatiese ontwikkeling, wat verhoogde groeitempo en n verkorte tyd tot bemarking beteken. Op soortgelyke wyse is dus gepostuleer dat triploiedie in perlemoen ook tot steriliteit kon lei. ‘n Triploiede induksie metode was ontwikkel deur Mathilde de Beer (2004) wat ‘n hoë persentasie triploidie in geinduseerde perlemoen opgelewer het. Die volgende logiese stap was om die groei van diploiede diere met die van triploiede diere te vergelyk om te bepaal of triploiedie wel ’n groei voordeel tot gevolg het met die oog op kommersiële toepassing. Deur van dieselfde tegnieke as De Beer (2004) gebruik te maak, is drie groepe, elk bestaande uit verwante diploiede en triploiede diere, geproduseer en ingeskryf aan n vergelykende groei proef. Die groepe was in drie verskillende seisoene geproduseer. Die hoof doelstelling van die proef was om groeitempo van diploiede en triploiede diere te vergelyk, asook om die invloed van seisoen op groei van diploide en triploide te bepaal. Twee groei eienskappe naamlik skulp lengte en liggaamsmassa is gemeet vanaf ‘n ouderdom van agt maande (wanneer die diere individueel gemerk kon word) tot ‘n ouderdom van 24 maande. Die algehele resultate het gedui op geen betekenisvolle verskil tussen die groei van triploiede en diploiede perlemoen tot op die ouderdom van twee jaar. Verskille het voorgekom in die groei tussen seisoene, maar daar kon nie bewys word dat die verskille die gevolg van seisoenale omgewingseffekte was nie. Diploiede en triploiede het dieselfde skulp lengte tot liggaamsmassa verhouding getoon tot op twee jaar ouderdom.

Thesis (MSc)--Stellenbosch University, 2008. / ENGLISH ABSTRACT: Haliotis midae, is the only commercially important species of the six abalone species found in South African coastal waters and has become a lucrative commercial commodity. Wild stocks of H. midae are, however, no longer commercially sustainable due to a combination of environmental factors and poaching. The solution to the crisis is artificial production systems in the form of abalone farms. An abalone enhancement programme was initiated in South Africa in 2006, funded by industry and government. This programme focuses on the elucidation of the abalone genome and genetic factors contributing to increased productivity, thereby aiding the commercial production of abalone. The aims of this study, the first of its kind concerning H. midae, were to develop AFLPbased markers (specifically fluorescent AFLP analysis); to monitor the segregation of these markers in a single full-sib family and to use the markers and additional microsatellite markers to generate the first preliminary linkage map for H. midae. Genomic DNA of sufficient quality and purity for fluorescent AFLP analysis was obtained from 3.5-month-old H. midae juveniles. Preliminary linkage maps were constructed using AFLP and microsatellite markers segregating in an F1 family following a pseudo-testcross mapping strategy. Twelve AFLP primer combinations, producing 573 segregating peaks, and 10 microsatellite markers were genotyped in the parents and 108 progeny of the mapping family. Of the 573 segregating AFLP peaks genotyped, 241 segregated in a 1:1 ratio and 332 in a 3:1 ratio. Of these AFLP markers, 90 segregated according to the expected 1:1 Mendelian ratio and 164 segregated according to the expected 3:1 Mendelian ratio at the P = 0.05 level and were used for linkage analysis. Of the 10 microsatellite markers genotyped, nine were informative for linkage mapping analysis. Preliminary male and female genetic linkage maps were developed using markers segregating in the female or male parent. A total of 12 and 10 linkage groups were detected for the female and male maps respectively. The female map covered 1473.5cM and consisted of 56 markers, and the male map covered 738.9cM consisting of 30 markers. Markers with segregation distortion were observed as previously reported in other abalone species and potential homology between one of the linkage groups of the male map and two of the linkage groups of the female map were identified using the 3:1 segregating AFLP markers. In conclusion, the genetic linkage map presented here, despite the fact that it has relatively low genome coverage and low marker density, forms an ideal starting point for more detailed study of the H. midae genome and will provide a scaffold for basic and applied studies in abalone. A high-density linkage map of H. midae should in future be developed with additional co-dominant molecular markers, such as microsatellites, to improve the transferability of the linkage map between different laboratories and among populations. A high-density linkage map will facilitate the mapping of QTL of commercially important traits (i.e. growth) and future MAS breeding programmes. / AFRIKAANSE OPSOMMING: Perlemoenspesie, Haliotis midae, is die enigste spesie van kommersiële belang van die ses wat in die kuswater van Suid-Afrika aangetref word en het ‘n winsgewende handelskommoditeit in Suid-Afrika geword. Die ontginning van natuurlike H. midae populasies is egter, as gevolg van ‘n kombinasie van omgewingsfaktore en stropery nie meer kommersieel volhoubaar nie. Die perlemoenkrisis kan die hoof gebied word deur kunsmatige produksiesisteme op perlemoenplase tot stand te bring. ‘n Perlemoen verbeteringsprogram is in 2006 in Suid-Afrika geïnisieer en word deur die industrie en regering befonds. Die program focus op die ontrafeling van die perlemoen genoom en die genetiese faktore wat bydrae tot verhoogde produksie. Sodanige inligting kan gebruik word om kommersiële perlemoenproduksie te bevorder. Die doel van hierdie studie, die eerste met H. midae, is om AFLP-gebaseerde merkers (spesifiek fluoresserende AFLP analise) te ontwikkel; die segregasie van hierdie merkers te monitor in ‘n enkel volledige verwante familie en die merkers en addisionele mikrosatelliet merkers te gebruik om die eerste voorlopige koppelingskaart vir H. midae te genereer. Genomiese DNS van genoegsame kwaliteit en suiwerheid vir fluoresserende AFLP analise is ge-ekstraeer uit 3.5-maand-oue H. midae individue. Voorlopige koppelingskaart is gekonstrueer deur van segregerende AFLP en mikrosatelliet merkers in ‘n F1 familie gebruik te maak deur ‘n pseudo-kruistoets karteringstrategie te volg. Twaalf AFLP inleier kombinasies, wat 573 segregerende fragmente geproduseer het, en 10 mikrosatelliet merkers is gegenotipeer in die ouers en 108 individue van die nageslag van die karteringsfamilie. Van die 573 segregerende AFLP merkers wat gegenotipeer is, het 241 in ‘n 1:1 verhouding en 332 in ‘n 3:1 verhouding gesegregeer. Van hierdie AFLP merkers, het 90 volgens die verwagte 1:1 Mendeliese verhouding en 164 volgens die 3:1 Mendeliese verhouding by die P = 0.05 gesegregeer vlak en is vir die koppelingsanalise gebruik. Van die 10 mikrosatelliet merkers gegenotipeer, was 9 informatief vir koppeling karteringsanalise. Voorlopige manlike en vroulike genetiese koppelingskaarte is ontwikkel met gebruik te maak van merkers wat in die manlike of vroulike ouer segregeer het. ‘n Totaal van 12 en 10 koppelingsgroepe is onderskeidelik in die vroulike en manlike karate gegenereer. Die vroulike kaart dek 1473.5cM and bestaan uit 56 merkers, terwyl die manlike kaart 738.9cM beslaan het met 30 merkers. Merkers wat segregasie distorsie toon is waargeneem soos voorheen in ander perlemoenspesies gerapporteer. Potensiële ooreenstemming tussen een van die koppelingsgroepe van die manlike kaart en twee van die koppelingsgroepe van die vroulike kaart is aangetoon deur van die 3:1 segregerende AFLP merkers gebruik te maak. Die genetiese koppelingskaarte verskaf wel ‘n relatiewe lae genoomdekking en ‘n lae merkerdigtheid, maar is ‘n ideale vertrekpunt vir meer gedetailleerde studie van die H. midae genoom en dien as ‘n raamwerk vir toekomstige basiese en toegepaste studies in perlemoennavorsing. ‘n Hoëdigtheid koppelingskaart van H. midae moet in die toekoms ontwikkel word met gebruik van bykomstige ko-dominante molekulêre merkers, soos mikrosatelliete. Dit sal die oordraagbaarheid van die koppelingskaart tussen verskillende laboratoria asook tussen populasies verbeter. ‘n Hoëdigtheid koppelingskaart sal die kartering van kwantitatiewe kenmerk loki (KKL) vir kommersieel belangrike kenmerke (onder andere groeikrag) en toekomstige merker bemiddelde seleksie (MBS) teelprogramme moontlik maak.