Thesis (MSc (Genetics))--University of Stellenbosch, 2009. / ENGLISH ABSTRACT: Oesophageal cancer (OC) is the sixth leading cause of cancer related deaths in the world with approximately 300 000 new cases reported each year. OC may be characterized into two forms with 90% of cases presenting as squamous-cell carcinoma (SCC) and the remaining 10% as adenocarcinoma (ADC). Several factors have been attributed to the development of OC, including oesphageal injury and/or irritation, chronic inflammation and excess iron associated with enhanced tumour growth. The HAMP gene codes for a 25 amino-acid protein found to be primarily expressed in the liver and crucial to regulation of bodily iron status. Defects occurring in the HAMP gene could therefore lead to the dysregulation of the gene, resulting in an iron overload status. Iron overload is a previously described risk factor in the development of various cancers, including OC, and therefore the aim of this study was to investigate whether dysregulation of the HAMP gene may be involved in the cancer phenotype exhibition. The study cohort comprised of 48 unrelated patients presenting with SCC and a control group of 51 healthy, unrelated population-matched individuals. Mutation detection techniques included polymerase chain reaction (PCR) amplification, heteroduplex single-stranded conformation polymorphism (HEX-SSCP) analysis and bi-directional semi-automated DNA sequencing analysis. Screening of the 5’ regulatory region (5’UTR) of the HAMP gene revealed one known (-582A/G) and two novel (-188C/T and -429G/T) variants with the -429G/T variant showing statistically significant reduction in expression in patients relative to controls. Iron parameters were correlated between patient and control cohorts, as well as for variant presence and absence within individuals. Luciferase reporter constructs were used to investigate the functional implications of the presence of a variant on HAMP gene expression, and how these results correlated to the iron parameter statistics obtained. Luciferase reporter assay results indicated the -188C/T and -429G/T variants to result in under-, and the -582A/G variant to result in over-expression at the basal level, relative to the respective wild-type sequence constructs. Correlation of the luciferase data with the iron parameter statistics, indicate the -429G/T variant to be coupled to significantly higher levels of ferritin and C-reactive protein (CRP) and significantly lower levels of serum-iron and transferrin when compared to individuals without the variant. Considering only the patient group, the presence of the -188C/T and -429G/T variants were coupled to significantly lower levels of transferrin in patients with either variant, compared to patients without. The variants found within the HAMP promoter region are therefore able to alter gene regulation to an extent where iron parameters deviate between healthy and OC afflicted individuals, and also between patients with and without a variant. This dysregulation in iron homeostasis may play a role in the development and/ or progression of OC. Characterisation of the 5’ UTR of the HAMP gene may contribute to linking iron regulation to the establishment of an effective screening program, facilitating the early detection of OC. / AFRIKAANSE OPSOMMING: Slukdermkanker (SK) is die sesde grootste oorsaak van kanker-verwante sterftes in die wêreld, met sowat 300 000 nuwe gevalle wat aangemeld word elke jaar. SK kan geklassifiseer word in twee vorme, waar 90% van die gevalle plaveisel-selkarsinoom (SSC) vorm en die oorblywende 10%, adenokarsinoom (ADC). Verskeie faktore word toegeskryf aan die ontwikkeling van SK, insluitend slukderm beserings en/ of irritasie, chroniese inflammasie en oormatige ystervlakke wat geassosieer word met verhoogde gewasgroei. Die HAMP geen kodeer vir 'n 25 aminosuur proteïen wat hoofsaaklik in die lewer uitgedruk word en noodsaaklik is vir die regulering van ystervlakke in die liggaam. Defekte wat in die HAMP geen voorkom kan dus die onreëlmatige regulering van die geen tot gevolg hê, wat lei tot yster-oorlading. Yster-oorlading is voorheen beskryf as ‘n risiko faktor in die ontwikkeling van verskillende vorme van kanker, insluitend SK en gevolglik was die doel van hierdie studie om te bepaal of die wanregulering van die HAMP geen betrokke mag wees by die uitdrukking van die kanker fenotipe. Die studiepopulasie het bestaan uit 48 onverwante pasiënte met SSC en ‘n kontrole-groep van 51 gesonde, onverwante soortgelyke individue. Die mutasie opsporingstegnieke wat gebruik is, het polimerase kettingreaksie (PKR) amplifisering, heterodupleks enkelstring-konformasie polimorfisme (HEX-SSCP) analise en bidireksionele semi-outomatiese DNS volgordebepaling-analise van die geïdentifiseerde variante ingesluit. Sifting van die 5’ regulerende area (5'UTR) van die HAMP geen het een bekende (-582A/G) en twee nuwe (-188C/T en -429G/T) variante opgelewer, met die -429G/T variant wat statisties beduidend onderdruk is in pasiënt uitdrukkings vlakke relatief tot 'n gesonde kontole-groep. Yster-parameters van alle pasiënt en kontole individue is gekorreleerd tussen pasiënt en kontrole groepe, sowel as vir teenwoordigheid of afwesigheid van variante in elke individu. Luciferase verklikker konstrukte is gebruik om die funksionele implikasies van die teenwoordigheid van ‘n variant op HAMP geenuitdrukking te ondersoek, en hierdie resultate te korreleer met yster-parameter statistieke wat verkry is. Luciferase verklikkertoetse dui aan dat die -188C/T en -429G/T variante tot verminderde, en die -582A/G variant lei tot die verhoogte uitdrukking op die basale vlak lei, relatief tot die onderskeie wilde-tipe konstukte. Korrelasie van die luciferase data met die yster-parameter statistieke, dui aan dat die -429G/T-variant gekoppel is aan aansienlik hoër vlakke van feritien en C-reaktiewe proteïen (CRP) en beduidend laer vlakke van serum-yster en transferrien in vergelyking is met individue sonder die variant. Met oorweging van slegs die pasiënt-groep, is die teenwoordigheid van die -188C/T en -429G/T variante beduidend gekoppel aan laer vlakke van transferrien in pasiënte met die variant, in vergelyking met pasiënte daarsonder. Variante binne die HAMP promotor is dus in staat om geenregulasie te verander tot so 'n mate dat die yster-parameters afwyk tussen gesonde en SK geaffekteerde individue, sowel as tussen pasiënte met en sonder ’n variant. Hierdie wanregulering in yster homeostase kan 'n rol speel in die ontwikkeling en/ of die progressie van SK. Karakterisering van die 5’ regulerende area van die HAMP geen kan grootliks bydra om ysterregulasie te verbind met die implementering van ‘n effektiewe siftingsprogram, en sodoende die vroeë opsporing van SK fasiliteer.

Thesis (MScAgric)--Stellenbosch University, 2011. / ENGLISH ABSTRACT: Research into markers for the detection of genetic diversity and cultivar identification has become an important component of the genetic improvement of crops. However, the incorporation of marker assisted selection (MAS) as a tool for the identification and characterization of breeding material has not been fully implemented in the breeding of spring triticale at Stellenbosch University’s Plant Breeding laboratory (SU-PBL). The present study served as a case study in order to achieve this. The first part of the study concerned the detection of genetic diversity in 101 newly sourced triticale cultivars, from a USDA germplasm bank, together with five local control cultivars, in order to identify possible crossing parents. Eight SSR markers, including five derived from rye and three from wheat, and five agronomic characteristics were used to assess diversity. In seedling screening the foreign cultivars showed resistance towards the stem rust isolate used, but were mostly susceptible to the leaf rust isolate. Out of the 8 SSR markers tested, 7 markers were polymorphic and revealed 140 alleles varying from 12 to 26 with an average of 17.5 alleles per locus. The observed polymorphic information content (PIC) value ranged from 0.39 to 0.88 with an average of 0.70, indicative of the good discriminatory ability of the SSR markers. The data revealed that the South African cultivars were genetically closely related to cultivars from the USA and Canada. The second part of the study focused on the introgression of a blue aleurone layer gene (Ba), carried by a wheat cultivar, ‘Cltr1202STR’, and purple pericarp genes (Pp1; Pp3) also carried by a wheat cultivar, ‘Amethyst’, into a triticale background. Unfortunately the introgression of the purple pericarp genes failed. Two lines containing the blue aleurone layer, 11T023 and 11T028, were however successfully created. Molecular typing of these lines with SSR markers were able to show that BC4F1 line 11T023 (Ba) B was genetically similar to the recurrent parent ‘Agbeacon’; and that the BC4F1 11T028 line (Ba) A was closest to the ‘US2007’ recurrent parent. The study illustrated that MAS was a reliable tool for detecting genetic diversity in newly sourced germplasm, and assisted in making a backcross breeding effort more effective. The data generated from MAS could therefore clearly assist in making the SU-PBL breeding program more effective by moving, better informed, decision making toward data based partly on the genotype, thereby minimizing the risks associated with purely phenotypic based decisions. / AFRIKAANSE OPSOMMING: Navorsing rondom die gebruik van merkers vir die bepaling van genetiese-diversiteit en kultivar identifikasie is ‘n toenemend belangriker komponent vir die genetiese verbetering van gewasse. Die inkorporering van merker-bemiddelde-seleksie (MBS) as gereedskap vir die identifikasie en karaktarisering van telingsmateriaal is nog nie ten volle geïmplimenteer in die lente korogtelingsprogram van die Stellenbosch Universiteit Planteteeltlaboratorium (SU-PTL). Die studie het gedien as gevallestudie ten einde dit te bereik. Die eerste gedeelte van die studie het gehandel oor die tipering van die genetiese diversiteit van ‘n 101 kultivars verkry vanaf ‘n USDA kiemplasmabank saam met 5 plaaslike kontroles. Dit was gedien ten einde moontlike kruisings-ouers te kon identifiseer. Agt SSR merkers, insluitend vyf afkomstig van rog en drie vanaf koring, asook vyf agronomiese kenmerke is aangewend om die materiaal se diversiteit te tipeer. Saailingtoetsing is ook gedoen en het aangetoon dat die meeste kultivars weerstandig was vir die stamroes-isolaat, maar nie die blaarroes-isolaat nie. Van die agt SSR merkers getoets het sewe getoon om polimorfies te wees en het ‘n 140 allele gegee wat gewissel het vanaf 12 tot 26 per lokus met ‘n gemiddeld van 17.5. Die waargenome polimorfiese inligtings inhoud (PII) waarde het gewissel vanaf 0.39 tot 0.88 met ‘n gemiddeld van 0.70. Die merkers kon dus suksesvol diskrimineer. Die data het aangetoon dat die Suid-Afrikaanse kultivars genetiese die naaste verwant was aan die kultivars afkomstig vanaf die VSA en Kanada. Die tweede gedeelte van die studie het gefokus op die introgressie van ‘n blou aleuron-laag geen (Ba), afkomstig vanaf die koringkultivar ‘Cltr1202STR’, en twee pers-perikarp gene (Pp1; Pp3), afkomstig vanaf die koringkultivar ‘Amethyst’, na ‘n korog agtergrond. Ongelukkig het die oordrag van die pers-perikarp gene gefaal. Twee lyne wat die blou aleuron- laag bevat, 11T023 en 11T028, is egter suksesvol geskep. Tipering van die lyne met die SSR merkers het aangetoon dat die BC4F1 lyn 11T023 (Ba) B genetiese baie na aan die herhalende ouers ‘Agbeacon’ is en dat die BC4F1 11T028 lyn (Ba) A nader is aan die herhalende ouer ‘US2007’. Die studie het dus geïllustreer dat MBS gebruik kan word as ‘n betroubare manier om genetiese diversiteit te bepaal en by te dra tot die sukses van ‘n terugkruisingsprogram. Die data wat dus voortspruit uit MBS kan dus help om die SU-PTL se telingsprogram te assisteer in die besluitnemingsproses tydens teling deur beter genotipe gebaseerde besluite te neem wat die riskio van fenotipe gebaseerde besluite kan help verminder.

Thesis (MScAgric)--Stellenbosch University, 2011. / ENGLISH ABSTRACT: Abalone production is the largest financial contributor to aquaculture in South Africa and practically all of the abalone produced is exported to Asia. This means that the product must be globally competitive and many technologies have been applied to this cause. One that specifically shows great promise for bivalve mollusc production is triploidy; more precisely, sterility due to the induction of aneuploidy. Under normal maturation, energy is diverted from somatic growth through sexual maturation, therefore inhibiting or retarding gametogenesis through a process such as aneuploidy is expected to increase growth and decrease the time to marketing. Two studies preceding this one investigated the induction of triploidy through hydrostatic shock (De Beer, 2004) and the comparative growth rate of triploid genotypes from 8 to 24 months, prior to the onset of sexual maturation (Schoonbee, 2008). During this comparative growth stage, no convincing statistical evidence of faster growth or of seasonal environmental effects could be obtained. It was recommended that growth between triploid and diploid variants be compared during the age period when sexual maturity becomes a factor to determine whether triploidy in Haliotis midae is a useful biotechnological tool to improve biological productivity and global competiveness of the abalone industry. The growth measured as shell length and wet weight in the period from 29 to 62 months showed a statistically significant difference in mean weight and mean length with diploids showing a superior growth rate compared to their triploid siblings. This difference of 1.99 mm and 5.13 g was however not perceived as being commercially significant. Important production parameters including canning yield percentage and gonadosomatic index were also measured during this trial. For both these parameters, the triploid genotype showed statistically and commercially significant improvement of 10.68% increased canning yield and 28.42% reduction in gonadosomatic index when compared to their diploid counterparts. Triploid abalone was found to be not completely sterile; gametes and even mature gonads were observed in some instances. Even though complete sterility was not achieved there appeared to be a retarded gonadosomatic development in triploid variants. The delay in sexual maturation, together with the improvement in canning yield, may justify triploidy’s commercial application, despite its reduced growth rate. / AFRIKAANSE OPSOMMING: Perlemoen produksie lewer die grootste finansiële bydra tot akwakultuur in Suid Afrika en feitlik al die Perlemoen word uitgevoer na Asië. Dit beteken dat die produk moet kompeteer op die wêreld mark en verskeie tegnologieë word reeds aangewend vir die spesifieke doel. Een so tegnologie wat potensiaal toon ten opsigte van akwakultuur produksie is triploïedie; meer spesifiek, sterieliteit veroorsaak deur aneuploïedie induksie. Onder normale volwassewording, word energie weggeneem van somatiese groei wanneer geslagsrypheid intree en daarom kan groeitempo verhoog word deur gametogenese te inhibeer of te vertraag deur ‘n proses soos aneuploïedie en, word korter tydperk benodig om bemarkingsgrootte te bereik. Twee voorafgaande studies het gehandel oor die induksie van triploïedie deur hidrostatiese druk skok (De Beer, 2004) en die vergelykende groeitempo van triploïede genotipes vanaf ouderdom 8 tot 24 maande (Schoonbee, 2008) alvorens geslagsrypheid intree. Tydens hierdie vergelykende groeifase kon geen statisties betekenisvolle aanduidings van vinniger groei of seisoenale omgewingseffekte aangetoon word nie. Die studie handel vervolgens oor die uitbreiding van die vergelykende groeistudies tussen triploïede en diploïede genotipes tot ouderdom van 62 maande wat die intrede van geslagsrypheid insluit, ten einde te bepaal of die induksie van triploïedie in Haliotis midae voordele inhou ten opsigte van produksiedoeltreffendheid en mededingendheid op wêreldmarkte. Groei gemeet in terme van skulplengte en lewende massa oor die tydperk van 29 tot 62 maande het statisties betekenisvolle verskille getoon in gemiddelde massa en lengte van diploïede genotipes bo die van triploïede verwante individue. Die verskille van 1.99 mm en 5.13 g kan egter nie as kommersieel betekenisvol beskou word nie. Belangrike produksie eienskappe waaronder persentasie opbrengs van eindproduk en die gonadosomatiese indeks is ook bepaal. Vir beide die produksie eienskappe het die triploïede genotipe statisties sowel as kommersieel betekenisvolle verbetering van 10.68% getoon vir opbrengs en 28.42% verlaging in gonadosomatiese indeks in vergelyking met die diploïede genotipe. Triploïede genotipes was nie volledig steriel nie, gegewe die aanwesigheid van gonades en gamete in sommige individue. Selfs al is totale steriliteit nie bereik nie, het dit wel voorgekom asof daar vertraging in gonadosomatiese ontwikkeling plaasgevind het in triploïede genotipes. Die vertraging in geslagsrypheid tesame met die verhoogde persentasie opbrengs van die eindproduk hou voordele in bo die andersins effense stadiger groei van triploïede genotipes.

Thesis (MSc (Genetics))--University of Stellenbosch, 2009. / Gene transfer from wild gras species to wheat is complicated by the simultaneous integration of large amounts of alien chromatin. The alien chromatin containing the target gene is inherited as a linkage block and the phenomenon is known as linkage drag. The degree of linkage drag depends on whether, and how readily, recombination occurs between the foreign and wheat chromatin. The S13 translocation line was developed by the department of Genetics, US. A cross was made between Chinese Spring and a leaf rust resistant Aegilops speltoides accession. Resistant backcross F1 was backcrossed to Chinese Spring and W84-17. S13 was selected from the backcross progeny and found to carry three rust resistance genes temporarily named LrS13, SrS13 and YrS13. Unfortunately, the resistance genes were completely linked to gametocidal (Gc) genes that were co-transferred from the wild parent. In wheat Gc genes cause reduced fertility, poor plant phenotype and hybrid necrosis. In order to use employ the rust resistance genes commercially they need to be separated from the Gc genes. At the onset of this study four putative shortened forms of the S13 translocation were provided. The four lines were identified in a homoeologous paring induction experiment (involving the test cross 04M127). This study aimed to achieve the following: (i) characterize the four recombinants with the use of molecular markers, (ii) use the knowledge gained to identify further recombinants in the 04M127 cross, (iii) identify the shortest (most useful) recombinant, and (iv) attempt to shorten the shortest recombinant form still further and thereby remove as many of the Gc genes as possible. In total, seven recombinants of the S13 translocation (04M127-1, -2, -3, -4, -7, -11 and -12; referred to as recombinant group A) were identified and characterised with microsatellite and SCAR markers. These recombinants have exchanged different amounts of foreign chromatin for wheat chromatin, but were still associated with Gc genes, showing hybrid necrosis and seed shrivelling. Some of the recombinants have lost the undesirable „brittle rachis‟ phenotype which occurs in Ae. speltoides and the S13 translocation line. In plants VII having this trait, the rachis spontaneously disarticulates after the third spikelet upon ripening of the ear. Recombinant 3 appeared to be least affected by Gc genes and was therefore used in further attempts to shorten the translocation. Recombinant 3 was crossed with wheat (W84-17) and resistant F1 (heterozygous for the translocation) were test crossed with Chinese Spring nullisomic 3A tetrasomic 3B/D plants. Thirty five resistant testcross F1 plants were identified (named recombinant group B). The resistant group B recombinants as well as nine susceptible test cross F1 (which also appeared to be recombinant) were characterised making use of microsatellites and a SCAR marker. From the results it appeared that each of the 35 resistant plants exchanged substantial amounts of Ae. speltoides chromatin for wheat chromatin. The species chromatin that remained (and which contains LrS13) is probably located either close to the 3AS telomere or within the proximal regions of 3AS and 3AL. A SCAR marker that has been developed specifically for the S13 translocation provided useful confirmation of the presence of Ae. speltoides chromatin in the 35 recombinants. If the SCAR marker proves to be tightly linked to LrS13 it may eventually be used for marker assisted selection of the resistance or it may be employed in continued attempts to reduce the amount of foreign chromatin. Seedling rust resistance tests showed that the recombinants have lost SrS13 and YrS1 during recombination. An attempt was also made to develop additional markers that specifically detect the translocation in order to further characterise the group B recombinants. Published information on Ae. speltoides specific repeated and transposon sequences were obtained and used for primer design. Unfortunately, no suitable markers could be found and the primers that were designed tended to amplify the same fragments in both the wheat and species genomes. DArT markers were also employed in an attempt to characterise the 35 group B recombinants and controls. The DArT results provided an independent verification of the results obtained with the microsatellite markers. The DArT results confirmed that the group B recombinants exchanged large amounts of species chromatin for wheat chromatin. Even though the 35 resistant group B recombinants have undergone extensive recombination they still show signs of residual Gc effects. It is believed these effects can be removed by continued backcrossing to wheat accompanied by selection against Gc symptoms. While the effects of Gc genes per se were not studied, their properties were reminiscent of those of transposable elements. Indications were that complex interactions involving the Gc genes themselves as well as genetic factors in the wheat genome may have a drastic effect on the selective survival of recombinant gametes.

MScAgric / Thesis (MSc (Genetics))--University of Stellenbosch, 2009. / ENGLISH ABSTRACT: In the Western Cape small grain cereals, triticale (×Triticosecale Wittmack ex A. Camus) in particular, appear to be among the most promising starch-carrying raw materials for the production of bio-ethanol. A core group of cultivars and lines from the Stellenbosch University Plant Breeding Laboratory spring triticale breeding programme were subjected to initial testing for the purpose of ethanol production. They underwent multi-location field-testing across six (season 2006–2007) and nine (season 2007–2008) locations representing the Western Cape cereal production area. Climatic conditions during the study were characterised as generally favourable, especially in the 2007 season. During the season, trials were visited in order to make in situ observations. Disease susceptibility was given specific attention. After harvesting, grain yield (kg.ha-1), test weight (kg.HL-1), total starch content in whole grain (%), amylose/amylopectin ratio, protein content (%), ethanol output (L.tonne-1) and ethanol yield (L.ha-1) were analysed. Near infra-red reflectance spectroscopy calibration models were developed for moisture and starch contents. The best calibration based on whole grain spectra for moisture content had RPD = 1.691, R2 = 0.657 and SEP = 0.271%, and for starch content RPD = 1.646, R2 = 0.634 and SEP = 1.356%. Calibrations developed from milled grain showed better results for moisture content RPD = 2.526, R2 = 0.843, SEP = 0.182%, and for starch content RPD = 1.741, R2 = 0.673, SEP = 1.277%. These calibrations are suitable for rough screening of samples. In the 2006 season, starch yield was highly positively correlated with grain yield (R2 = 0.988, P <0.001). Both starch yield and grain yield were positively correlated with days to heading (R2 = 0.533 and R2 = 0.556, respectively; P <0.001). The 2007 season was characterised by a generally higher starch yield (2952– 3142kg.ha-1, 95%CI) compared to the 2006 season (2077–2315kg.ha-1, 95%CI). Starch yield was strongly positively correlated with grain yield (R2 = 0.975, P <0.001). Test weight demonstrated weak positive correlation with ethanol yield (R2 = 0.238, P <0.01) and grain yield (R2 = 0.279, P <0.001). Mean ethanol output ranged between 466–477L.tonne-1 at the 95%CI. Ethanol output was demonstrated to be more dependent on starch and other polysaccharides accessibility to enzymatic digestion than on the total starch content as such. The best lines for ethanol output in the 2007 season were G2, D3 and H2 for the Swartland region, and D3, G2 and D1 for the Overberg region. The best triticale lines under investigation showed their potential from a biological point of view to be a suitable crop for ethanol production in the Western Cape, with the achieved ethanol yield ranging between 2446–2625L.ha-1 at the 95%CI. For the Swartland region the best genotypes for ethanol yield were D1, H1 and D2, and for the Overberg H1 and G2. The 23 best lines were selected from the elite and senior blocks, and then used for the establishment of a recurrent massselection pre-breeding block. / AFRIKAANSE OPSOMMING: In die Wes-Kaap is kleingrane, meer spesifiek korog (×Triticosecale Wittmack ex A. Camus), van die mees belowende styseldraende rou-materiale vir die produksie van bio-etanol. ‘n Kern versameling van kultivars en telerslyne van die Universiteit van Stellenbosch se Planteteeltlaboratorium se lente korogteeltprogram is blootgestel aan aanvanklike toetsing met die doel om etanol produksie te meet. Die materiaal het veldtoetsing ondergaan oor verskeie lokaliteite gedurende die 2006–2007 (ses lokaliteite) en 2007–2008 (nege lokaliteite) seisoene wat verteenwoordigend was van die Wes-Kaapse produksie gebied. Klimaatstoestande gedurende die studie kan beskryf word as gunstig, veral gedurende die 2007 seisoen. Gedurende die groeiseisoen is proeflokaliteite gereeld besoek ten einde in situ observasies te kon maak, siektevatbaarheid het veral aandag geniet. Na die oes van proewe was graanopbrengs (kg.ha-1), hektolitermassa (kg.HL-1), totale-styselinhoud in heelgraan (%), amilose/amilopektien-verhouding, proteïeninhoud (%), etanolopbrengs (L.ton-1) en etanolopbrengs per hektaar (L.ha-1) gemeet. Naby-infrarooispektroskopie kalibrasies was ontwikkel vir vog- en styselinhoud. Die beste kalibrasies vir heelgraan voginhoud het ‘n RDP = 1.691, R2 = 0.657 en SEP = 0.271% en vir styselinhoud RPD = 1.646, R2 = 0.634 en SEP = 1.356% opgelewer. Die kalibrasies gebaseer op meel was aansienlik beter vir voginhoud RPD = 2.526, R2 = 0.843 en SEP = 0.182%, sowel as vir styselinhoud RPD = 1.741, R2 = 0.673 en SEP = 1.277%. Die kalibrasies is bruikbaar vir aanvanklike sifting van monsters. 5 Gedurende die 2006 seisoen het styselinhoud en graanopbrangs ‘n baie hoë korrelasie (R2 = 0.988, P <0.001) getoon. Beide stysel- en graanopbrengs was positief gekorreleerd met dae tot aar (R2 = 0.533 en R2 = 0.556; P <0.001). Die 2007 seisoen is gekenmerk deur ‘n hoër styselopbrengs (2952– 3142kg.ha-1, 95%VI) teenoor die 2006 seisoen (2077–2315kg.ha-1, 95%VI). Styselopbrengs was positief gekorreleerd met graanopbrengs (R2 = 0.975, P <0.001). Hektolitermassa het swak korrelasie getoon met etanolopbrengs (R2 = 0.238, P <0.01) en graanopbrengs (R2 = 0.279, P <0.01). Gemiddelde etanolopbrengs het gewissel tussen 466–477L.ton-1 by 95%VI. Data het aangedui dat etanolopbrengs meer aangewese is op stysel en ander polisakkariedverbindings se ensiematiese toeganklikheid eerder as totale stysel aanwesig. Die beste lyne wat etanolopbrangs betref in 2007 was G2, D3 en H2 vir die Swartland en D3, G2 en D1 vir die Overberg. Van die koroglyne wat deel was van die ondersoek het goeie potensiaal getoon, uit ‘n suiwer biologiese oogpunt, as gewas vir die produksie van etanol in die Wes-Kaap met ‘n gerealiseerde etanolopbrengs in die omgewing van 2446-2625L.ha-1 by 95%VI. In die Swartland was die beste genotipes D1, H1 en D2 en in die Overberg H1 en G2. Die beste 23 lyne is geselekteer uit die elite en senior telingsblokke en aangewend in die vestiging van ‘n herhalende-seleksie voortelingsblok.

The aim of this study was to analyse testcross-material that was generated during a homoeologous pairing-induction experiment. Absence of the homoeologous pairing suppressor gene, Ph1, was employed to induce meiotic pairing between the Lr59 translocation (Aegilops peregrina) and 1AL of normal wheat. The study aimed to characterize the test-cross plants derived from this experiment and to identify recombinants which retained the least amount of species chromatin but which still contained the Lr59 gene. The test-cross F1 population, 07M5 (total 635 plants), was screened for Lr59 resistance by inoculating seedlings with the leaf rust pathotype, UVPrt8. The 168 resistant plants were characterized with molecular markers in order to identify recombinants. The data were used to construct a physical map which showed the relative sizes of the recombinants and which could be used to identify those recombinants which contained the least amount of residual species chromatin. Microsatellite (Xcfa2219, Xbarc83 and Xgwm164) and SCAR (S15T3) analysis was used for the initial identification of recombinants. The results showed that 152 of the 168 resistant plants were recombinants for the four loci; that eight of the remaining 16 plants represented non-recombinant, wild species-types and that the last eight plants represented the wheat parental-types which were resistant (and thus, also recombinants). This extremely high recombination frequency can largely be attributed to strong segregation distortion that was evident in the cross. It is also possible that the translocation segment could derive from the S genome rather than the U genome of Ae. peregrina. The S genome is closer related to the wheat genomes than the U genome and may be more prone to recombination. With the use of the microsatellite and SCAR data, a physical map was constructed which showed the relative location of the Lr59 gene on the translocation. It appeared that the eight shortest recombinants retained terminal species chromatin. In an attempt to characterize the eight recombinants, additional marker loci had to be identified within that region. RAPD, iv AFLP and DArT markers were investigated for this purpose. RAPD analyses did not produce any useful markers. AFLP and DArT analyses did identify useful markers with which the eight recombinants could be screened. The data showed which recombinants probably retained the least amount of species chromatin. Seeing that AFLP and DArT markers are anonymous and that the distances between marker loci are unknown, it is not possible to say which recombinant is the shortest and consequently it will be nessecary to also evaluate the group of eight recombinants agronomically in order to identify the most useful ones. The results showed that multiple cross-overs apparently occured on both sides of Lr59. Multiple cross-overs are higly unlikely in material of this nature, therefore it was speculated that the observation resulted from incomplete synteny between the telomeric areas of the translocation and 1AL. A structural difference between the two chromosome regions might have given rise to abnormal meiotic pairing structures and thus unexpected gamete genotypes. Each of the eight recombinants did express one or more of the Ae. peregrina derived AFLP loci which can in future be verified for use as a marker for marker assisted selection. The study succeeded in identifying a number of potentially useful recombinants which contain the Lr59 resistance. It would, however, be risky to select only one of the shortest recombinants for further development on the basis of the present knowledge as some recombinants may contain genetic abnormalities which resulted from reduced synteny in the Lr59 region. It would therefore be wise to further evaluate all eight recombinants before the best one is selected for agronomic use.

Thesis (MSc)--Stellenbosch University, 2012. / ENGLISH ABSTRACT: Biopolymers exhibit the required material properties to replace conventional, non-biodegradable, petroleum-based polymer products. They have a closed carbon cycle, making them carbon neutral and environmentally friendly. Biopolymers are produced from non-toxic substrates during in vivo enzymatic reactions. Biosynthesis of the most commercially important biopolymers is too complex to be reproduced in in vitro reactions. Identification of the genes responsible for their biosynthesis has been under investigation, with some pathways already elucidated. The genes involved in the biosynthesis of these polymers have been targeted for genetic manipulation to increase productivity, as well as create tailor-made polymers. Novel biopolymers and the genes responsible for their synthesis are of interest for their potential commercial applications. Bacteria produce a wide range of biopolymers and are being implemented as the bio-factories for biopolymer production. They are capable of utilising easily accessible and renewable carbon sources such as sucrose for polymer biosynthesis. Bacteria thus allow for economical production of these environmentally beneficial polymers. In this study, the gene responsible for the production of an unknown biopolymer from an unknown bacterium was identified. The biopolymer producing bacteria were grown on media enriched with sucrose as carbon source, during an expression library screening in a previous study. Expression library technology was used to search for the gene and it was identified as a 424 amino acid levansucrase which had a 100% homology to Leuconostoc mesenteroides M1FT levansucrase (AAT81165.1). Biopolymer analysis revealed that the biopolymer was a levan, a polysaccharide consisting of only fructose molecules with a molecular weight of ± 5 kDa. Analysis of a 516 bp fragment of the 16S rRNA determined that the unknown bacteria were a Pseudomonas species. / AFRIKAANSE OPSOMMING: Bio-polimere besit noodsaaklike materiële eienskappe wat toelaat dat dit konvensionele, nie bio-afbreekbare, petroleum-gebasseerde polimeer produkte kan vervang. Hulle het n geslote koolstof kringloop en is dus koolstof neutraal en omgewingsvriendelik. Bio-polimere word vervaardig van nie-toksiese substrate, gedurende ensiematiese reaksies in vivo. Die belangrikste kommersiële bio-polimere se ensiematiese produksie is te kompleks om in ʼn in vitro reaksie te herproduseer. Ondersoeke tot die identifikasie van die gene wat verantwoordelik is vir die produksie van die polimere is onderweg, en sommige produksie paaie is reeds bekend. Die bekende gene word geteiken vir genetiese manipulasie om hulle produktiwiteit te vermeerder en om unieke polimere te produseer. Unieke bio-polimere en die gene wat vir hul produksie verantwoordelik is, is van belang vir hulle potentiële implimentering in komersiële toepassings. Bakteria produseer ʼn verskeidenheid bio-polimere en word as die bio-fabrieke vir polimeerproduksie geimplimenteer. Hulle kan maklik bekombare koolstofbronne, soos sukrose, gebruik om bio-polimere te produseer. Bakteria laat dus die ekonomiese produksie van hierdie omgewingsvriendelike polimere toe. In hierdie studie word die geen wat verantwoordelik is vir die produksie van ʼn onbekende bio-polimeer van ʼn onbekende bakteria, geidentifiseer. Die bakteria was gevind op media, wat verryk was met sukrose as koolstofbron, tydens ʼn vorige studie, waartydens ʼn uitdrukkingsbiblioteek gesif was op hierdie media. Uitdrukkingsbiblioteek tegnologie was gebruik om die geen te vind. Die geen was geidentifiseer as ʼn 424 aminosuur, homo-fruktose-polimeer produseerende geen, ʼn “levansucrase”. Die geen het ʼn 100% homologie met die M1FT “levansucrase” geen (AAT81165.1) van Leuconostoc mesenteroides gehad. Analise van die bio-polimeer het bepaal dat die polimeer ʼn polisakkaried was, wat slegs uit fruktose molekules bestaan het. Die molekulêre gewig van die polimeer was ± 5 kDa. Analise van ʼn 516 bp fragment van die 16S rRNS het bepaal dat die bakteria van die Pseudomonas spesie afkomstig was.

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.

Thesis (MSc)--Stellenbosch University, 2012. / ENGLISH ABSTRACT: The decline of Haliotis midae (perlemoen) populations together with the ensuing collapse of commercial abalone fisheries in South Africa have shifted the responsibility to abalone farms to meet the demand for perlemoen. Attention has recently turned to the genetic enhancement of cultured abalone in order for the farms to remain competitive in the international aquaculture market. To develop a successful breeding programme it is imperative to draw on a good foundation of high levels of genetic diversity and to successfully maintain these levels in order to create an enhanced strain of cultured abalone. A Performance Recording Scheme (PRS) was established as the first breeding programme for Haliotis midae to utilise molecular tools. This programme was aimed at enhancing the growth rate of abalone in order to shorten the production times on farms. The current study made use of 12 species-specific microsatellite markers to assign parentage to a group of faster-growing PRS animals, as selected by the abalone farms, in order to select a diverse on-farm generation of broodstock. Additionally, the influence of standard selection practises on the genetic diversity of a population compared to genotypic selection was investigated. This data was also used to study the differentiation and levels of genetic diversities within and between cultured and wild populations. Selection based on genotypic traits successfully retained genetic diversity while some diversity was lost in phenotypically selected populations. These phenotypic populations differed significantly from each other and wild populations, while the genotypic populations were similar in genetic composition to each other and wild populations of the West coast. The broodstock populations used in the PRS spawning event were representative of the wild populations from where they were sourced, with no significant differentiation between the broodstock and West coast population. When these broodstock populations were compared to their corresponding offspring populations, only two populations displayed a significant loss in diversity; although all of the offspring populations showed significant differentiation with their corresponding broodstock populations. This was attributed to the differential contribution of broodstock and the effect of artificial selection. It was established that the cultured populations of the participating abalone farms should be used with caution in ranching and reseeding programmes. These populations differed significantly from both the East and West coast wild populations. This study concluded that it is possible to retain genetic diversity by selecting breeding animals based on genotypic traits. The loss of diversity in some cultured populations and significant differentiation from the wild populations indicate that animals are exposed to different selection pressures in the cultured environment. The results found in this study highlight the need for the effective management of hatchery practices and the genetic monitoring of the breeding animals. / AFRIKAANSE OPSOMMING: Die afname in Haliotis midae (perlemoen) populasies en die daaropvolgende ineenstorting van die kommersiële perlemoen bedryf in Suid-Afrika het die verantwoordelikheid om in die aanvraag na perlemoen te voorsien, na perlemoen plase verskuif. Die genetiese verbetering van verboude perlemoen geniet tans aandag in ‘n poging om kompeterend te bly in die internasionale mark. Dit is noodsaaklik vir die sukses van ‘n broeiprogram om gebruik te maak van ‘n goeie genetiese basis met hoë vlakke van genetiese diversiteit en die suksesvolle behoud van die vlakke om so ‘n verbeterde lyn te skep. ‘n Groeiprestasie aanteken stelsel [Performance Recording Scheme (PRS)] is gestig as die eerste broeiprogram vir Haliotis midae wat gebruik maak van molekulêre tegnieke. Die doel van hierdie program was om die groeitempo van verboude perlemoen te verbeter om produksie tye te verkort. Die huidige studie het gebruik gemaak van 12 spesie-spesifieke mikrosatelliet merkers om ouerskap toe te ken aan ‘n groep vinnig-groeiende PRS-diere, soos geselekteer deur die perlemoen plase, om ‘n diverse generasie gekultiveerde diere te selekteer wat as broeidiere kan dien. Die invloed van standaard seleksie metodes op die genetiese diversitiet van ‘n populasie in vergelyking met genotipiese seleksie is ook ondersoek. Die ouerskap data is ook gebruik om differensiasie en vlakke van genetiese diversiteit tussen verboude perlemoene en wilde populasies vas te stel. Seleksie gebasseer op genetiese eienskappe het daarin geslaag om genetiese diversiteit te behou, terwyl diversiteit verlore gegaan het in die fenotipies geselekteerde populasies. Hierdie fenotipiese populasies het ook beduidend met mekaar sowel as met die wilde populasies verskil, terwyl genotipiese populasies soortgelyk was in hul genetiese samestelling en nie van die wilde populasies van die Weskus verskil het nie. Die broeidiere wat in die PRS broeiprogram gebruik is, was verteenwoordigend van die wilde populasies vanwaar hulle oorspronlik gekom het, met geen beduidende differensiasie tussen die broeidiere en die Wes kus populasies nie. Met die vergelyking van die broeidiere en hul ooreenstemmende nageslag, het dit geblyk dat slegs twee populasies ‘n beduidende verlies aan genetiese diversiteit getoon het, alhoewel al die nageslag beduidende populasie differensiasie met hul ouers getoon het. Hierdie bevindinge is toegeskryf aan oneweredige bydraes van die broeidiere tydens gameetvrystelling en die invloed van kunsmatige seleksie. Hierdie studie het ook vasgestel dat die verboude perlemoen populasies met sorg gebruik moet word om wilde populasies te herstel, aangesien hierdie populasies beduidend verskil het van wilde populasies van beide die Oos en Wes-kus. Hierdie studie het gevind dat dit moontlik is om genetiese diversiteit te behou deur diere te selekteer op grond van genotipiese eienskappe. Die verlies van diversiteit in sommige van die verboude perlemoen populasies en die beduidende verskil met die wilde populasies dui daarop dat diere in die gekultiveerde omgewing blootgestel word aan verskillende tipes seleksiedruk. Hierdie bevindinge beklemtoon die belang vir effektiewe bestuur van broeiery praktyke en genetiese monitering van broeidiere.

Thesis (MSc)--Stellenbosch University, 2012. / ENGLISH ABSTRACT: Haliotis midae, or more commonly known as Perlemoen, is an abalone species found along the coast of South Africa. It is the only cultured abalone species in South Africa and has a high demand abroad. Due to its popularity as a seafood delicacy, illegal harvesting has taken its toll on Perlemoen numbers. This increases the need for sustainable farming efforts and efficient implementation of law enforcement practices against poachers. Abalone farms make use of a limited number of broodstock for breeding, so it is necessary to ensure that genetic effects such as inbreeding and bottlenecks do not interfere with the viability of the offspring. Research that focuses on the genetics of Perlemoen will greatly aid the farms to continue sustainable production of this species as well as enhance their breeding efficiency. This study focuses on the construction of a linkage map for H. midae that will allow the future identification of markers associated with genes important to production, such as growth and disease resistance. Identification of these genes will allow breeders to select genetically superior abalone that will be used for breeding programmes in which the phenotype of the offspring will be enhanced. For the construction of a linkage map it is necessary to have enough informative markers for mapping. In this study, gene-linked microsatellite markers were developed by screening a contig assembly of H. midae’s transcriptome. Ninety-eight primer pairs could be developed from the contigs and 60 loci produced amplification products. Twenty-six microsatellites were found to be polymorphic (27% success rate). In addition to these markers, 239 previously developed microsatellites and 48 gene-linked SNPs were used to develop sex-average and sex-specific linkage maps in four full-sib families consisting of approximately 100 offspring each. Of these markers 99 were informative in family DS1 (31% success rate), 81 in family DS2 (26%), 77 in family DS5 (24%) and 71 in family DS6 (23%). These markers were used for linkage analysis (LOD>3). The average number of linkage groups for the sex-average maps ranged from 17-19. The average genome length for these maps ranged from 700cM to 1100cM with an average marker spacing of 8cM. The sex-specific maps’ linkage groups ranged from 13-17 with an average genome length of 600cM to 1500cM. The average marker spacing was approximately 16cM. The integrated map was constructed by merging the sex-average maps. This map contained 25 linkage groups with an average genome length calculation of 1700cM and an average marker spacing of 9.3cM. The linkage maps created in this study are the first to utilize SNPs in H. midae. Further incorporation of SNPs into linkage maps will enhance the density. The maps created in this study are of medium-density (65%) and provide a link to the development of high-density linkage maps to facilitate associations of phenotypic traits to certain markers, to so that QTL mapping can be performed. This information can be used for marker-assisted selection to produce genetically superior abalone. / AFRIKAANSE OPSOMMING: Haliotis midae, of meer algemeen bekend as Perlemoen, is 'n klipkous spesie wat langs die kus van Suid-Afrika voorkom. Dit is die enigste gekweekte klipkous spesie in Suid-Afrika en het 'n hoë aanvraag in die buiteland. As gevolg van sy gewildheid as 'n seekos lekkerny, het onwettige stropery sy tol geneem op Perlemoen getalle. Hierdie verhoog die behoefte vir volhoubare boerdery pogings en doeltreffende implementering van wetstoepassing teen stropers. Perlemoenplase maak gebruik van 'n beperkte aantal broeidiere vir teling, dus is dit nodig om te verseker dat genetiese effekte soos inteling en genetiese bottelnekke nie inmeng met die lewensvatbaarheid van die nageslag nie. Navorsing wat fokus op die genetika van Perlemoen sal grootliks die plase steun om die volhoubare produksie van hierdie spesie voort te sit, sowel as hul teling doeltreffendheid te verbeter. Hierdie studie fokus op die ontwikkeling van 'n genetiese koppelingskaart vir H. midae, wat die toekomstige identifisering van die merkers wat verband hou met die gene wat belangrik is vir die produksie, soos groei en weerstand teen siektes sal verbeter. Identifisering van hierdie gene sal toelaat dat telers genetiese voortreflike Perlemoen kan kies vir teelprogramme waartydens die fenotipe van die nageslag sal verbeter word. Vir die ontwikkeling van 'n genetiese koppelingskaart is dit nodig om genoeg informatiewe merkers vir die kartering te hê. In hierdie studie, is geen-gekoppelde mikrosatelliet-merkers ontwikkel deur ‘contig’ data van H. midae se transkriptoom te ondersoek. Agt en negentig inleier pare kon ontwikkel word uit die ‘contigs’ en 60 loki kon ‘n amplifiseringsproduk lewer. Ses-en-twintig mikrosatelliete was polimorfies (27% suksessyfer). Bykomend tot hierdie ontwikkelde merkers is 239 voorheen ontwikkelde mikrosatelliete en 48 geen-gekoppelde SNPs gebruik om geslagsgemiddelde en geslagspesifieke koppelingskaarte in vier volsib families, wat uit ongeveer 100 nageslag elk bestaan, te ontwikkel. Van hierdie merkers was 99 informatief in familie DS1 (31%), 81 in die familie DS2 (26%), 77 in die familie DS5 (24%) en 71 in die familie DS6 (23%). Hierdie merkers is gebruik vir 'n koppelingsanalise (LOD>3). Die gemiddelde aantal koppelingsgroepe vir die geslagsgemiddelde kaarte het gewissel van 17-19. Die gemiddelde genoom lengte vir hierdie kaarte het gewissel van 700cM tot 1100cM met 'n gemiddelde merker spasiëring van 8cm. Die koppelingsgroepe van die geslagspesifieke kaarte het gewissel van 13-17 met 'n gemiddelde genoom lengte van 600cM tot 1500cM. Die gemiddelde merker spasiëring was ongeveer 16cm. Die geïntegreerde kaart is saamgestel deur die samesmelting van die geslagsgemiddelde kaarte. Die kaart toon 25 koppelingsgroepe met 'n gemiddelde berekende genoom lengte van 1700cM en' n gemiddelde merker spasiëring van 9.3cM. Die genetiese koppelingskaarte wat in hierdie studie ontwikkel is, is die eerste om SNPs te gebruik in H. midae. Verdere insluiting van SNPs in koppelingskaarte sal die digtheid verhoog. Die kaarte wat in hierdie studie ontwikkel is, is van medium digtheid (65%) en bied 'n stap nader aan die ontwikkeling van hoë digtheid koppelingskaarte om fenotipiese eienskappe met sekere merkers te assosieer, vir kwantitatiewe kenmerk lokus kartering. Hierdie inligting kan gebruik word vir merker bemiddelde seleksie om geneties verbeterde Perlemoen te produseer.