Dissertation (PhD)--University of Stellenbosch, 2004. / ENGLISH ABSTRACT: Understanding the complex nature of grapevine molecular biology is of great importance for
viticulturists. Progress in the elucidation of key events on a genetic level could provide further
insight into the underlying cues responsible for the precise control of physiological and
metabolic changes during a specific condition such as fruit development. The use and analysis
of molecular ‘tools’, such as promoters controlling the site and level of gene activity, could
assist in the understanding of grapevine biology and serve as a platform for the future design
and development of recombinant DNA protocols and strategies for Vitis vinifera L.
A high-throughput gene expression system, cDNA-AFLPs, was successfully used to analyse
large-scale transcriptional activity during berry ripening. Candidate cDNA fragments were
selected on the basis of desired expression patterns and/or known gene function for subsequent
promoter isolation. From three candidate cDNAs selected, the promoter of a gene encoding
vacuolar pyrophosphatase (V-PPase) was isolated for computational and comparative analyses.
Promoter activity was evaluated on a transient level using the green fluorescent protein (GFP)
reporter gene. Comparative integration has allowed for putative correlation of cis-elements,
acting as receptors within promoter regions, to regulate V-PPase gene expression in response to
development, environmental stress and tissue-specificity.
In this study, integration of genetic data have advanced the understanding and transcriptional
role of a key enzyme (V-PPase) during grape ripening. Although never a replacement for
experimental verification, this integrative strategy of combining gene expression profiles with
bioinformatics and regulatory data will greatly assist in further elucidation of various other key
components and regulatory cues associated with grapevine molecular biology. This study has
allowed us to use molecular tools that could assist in gaining further insight into genetic
complexities and could serve as a platform for a more refined genetic manipulation strategy in
Vitis vinifera L. / AFRIKAANSE OPSOMMING: Begrip van die komplekse aard van wingerd molekulêre biologie is van groot belang vir wingerdkundiges. Vooruitgang in die begrip van belangrike gebeurtenisse op ń genetiese vlak behoort verdere insig in die onderliggende instruksies vir die noukeurige beheer van fisiologiese en metaboliese veranderinge tydens ń spesifieke kondisie soos vrug rypwording te bevorder. Die gebruik en analise van molekulêre ‘instrumente’ soos promoters, wat die posisie en vlak van geen aktiwiteit beheer, kan bydra tot n beter begrip van wingerd biologie en sodoende dien as ń platform vir die toekomstige ontwerp en ontwikkeling van rekombinante DNS (deoksiribonukleiensuur) protokolle en strategieë vir Vitis vinifera L. ń Hoë-kapasiteit geen uitdrukkings sisteem, nl. kDNS-AFLPs (komplementêre deoksiribonukleiensuur- geamplifiseerde fragment lengte polimorfisme), is suksesvol gebruik vir die analise van grootskaalse transkripsionele aktiwiteit tydens druif rypwording. Kandidaat kDNS fragmente is geselekteer, gebaseer op verlangde uitdrukkings-patrone en/of bekende geen funksie vir daaropvolgende promoter isolering. Van drie geselekteerde kandidaat kDNS fragmente, is die promoter van ń geen wat vakuolêre pirofosfatase (V-PPase) kodeer geïsoleer vir rekenaar- en vergelykende analise. Promoter aktiwiteit is op ń nie-stabiele vlak deur die gebruik van ń groen-fluoresserende proteien (GFP) verklikker geen geëvalueer. Vergelykende integrering het dit moontlik gemaak om veronderstelde korrelasies van cis-elemente, wat as reseptore binne ń promoter area dien, en die regulering van V-PPase geen uitdrukking, in reaksie tot ontwikkeling, omgewings stres en weefsel-spesifisiteit, te maak. Tydens hierdie studie, het die integrering van genetiese data gehelp om die transkripsionele rol van ń belangrike ensiem (V-PPase) tydens druif rypwording beter te verstaan. Alhoewel dit nooit ń plaasvervanger vir eksperimentele bewyse sal wees nie, kan hierdie gëintegreerde strategie, wat die kombinasie van geen-uitdrukkingsprofiele met bioinformatika en regulatoriese data behels, grootliks bydra om verskeie ander belangrike komponente en regulatorieseaanwysings geassosieërd met wingerd molekulêre biologie te ontrafel. Hierdie studie het verdere insig in genetiese kompleksiteite verleen, en kan nou dien as ń platform vir ń meer presiese genetiese manipulering strategie in Vitis vinifera L.
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:sun/oai:scholar.sun.ac.za:10019.1/16078 |
| Date | 04 1900 |
| Creators | Venter, Mauritz |
| Contributors | Botha, Frederik C., Potier, Bernard, University of Stellenbosch. Faculty of Science. Dept. of Botany and Zoology. |
| Publisher | Stellenbosch : University of Stellenbosch |
| Source Sets | South African National ETD Portal |
| Language | en_ZA |
| Detected Language | Unknown |
| Type | Thesis |
| Format | xvi, 97 leaves : ill. (some col.) |
| Rights | University of Stellenbosch |
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