Potato is a major part of the human diet in many countries of the world, providing substantial levels of carbohydrate, protein, and vitamins. This study examined the tuber protein content. In the first part of the research, total soluble protein (TSP) and patatin concentration were determined in periderm, cortex, and pith, in tubers of 20 important potato cultivars. TSP concentration was greater in periderm and lesser in cortex and pith tissues. Patatin was present in all tuber tissues but with the opposite pattern, less in periderm and greater in cortex and pith tissues. For intercultivar comparisons, a means of converting the specific tissue-based TSP and patatin data (dry weight) into a uniform weight whole tuber basis was developed. This relied on conversion factor values that were generated from percent weight tissue proportion and percent dry matter for each tissue layer. Cultivars with relatively more or less TSP and patatin in each tissue layer, and on a whole tuber basis, were identified. In the second part of the study, disassembly of chimeral (Russet Burbank) and putatively chimeral (Alpha, Bintje, Red Gold) tubers into their component genotypes was evaluated as a strategy for the production of intraclones with altered protein content. Explants were selected from tissue with greater or lesser protein levels and somatic embryogenesis was used to produce regenerants from each tissue source. Russeting was used as a phenotypic marker and TSP as a biochemical marker. Russet Burbank was confirmed as a periclinal chimera, although chimeral instability was evident, since some non-chimeral regenerants showed displacement of LI tunic cells with the russeting mutation into the pith. Red Gold was "uncovered" as an LII periclinal chimera (Red-Gold-Red). The value of chimeral disassembly in explaining an important component of somatic variation was clearly seen with this cultivar. The inconsistent TSP distribution in Russet Burbank intraclones proved that TSP was not distributed in a periclinal chimeral manner, as initially hypothesized. However, there was clear variation in protein content in the tubers of non-chimeral regenerants. Periclinal chimeral disassembly and somatic embryogenesis are potentially useful technologies for the production of improved intraclones of potato.

No description available.

Thesis (M.Sc.)(Plant Biotechnology))--University of Pretoria, 2002. / Summaries in Afrikaans and English. Includes bibliographical references.

Potato leafroll virus (PLRV) is one of the most destructive potato viruses in South Africa. In order to establish resistance against PLRV in commercial potato cultivars, the coat protein (CP) gene of the virus was previously isolated, cloned and subcloned into the plant expression vector pBI121 in both the sense and antisense orientations (BURGER, unpublished results). The pBI121 constructs containing the PLRV-CP gene were subsequently transferred to Agrobacterium tumefaciens LBA 4404 in a triparental mating process with the helper plasmid pRK2013. Two A. tumefaciens- mediated transformation methods for potatoes were investigated, viz. vacuum infiltration and leaf disk transformation. In addition, optimal transformation and regeneration conditions were identified for potato cultivars Late Harvest and BP[1] In total, 27 transgenic potato lines containing the PLRV-CP, β-glucoronidase (GUS) and nptII (neomycin phosphotransferase II) trans genes were generated under kanamycin selection. Transgenic plants grown in the glasshouse appeared to be phenotypically normal, and no differences in ploidy level in comparison to non-transformed plants could be established. Stable transgene insertion into the genome of the transgenic plants was verified using PCR and Southern blot analysis. Expression of the GUS transgene was investigated using a fluorometric assay (JEFFERSON et al. 1987), and it was found that orientation of the inserted PLRV-CP gene upstream from the GUS gene had a direct influence on the levels of GUS expression. The expression of the PLRV-CP gene was analysed using DAS-ELISA and immunoblot detection. Coat protein could not be detected in either assay. RNA dot blots were used successfully to show PLRV-CP expression in transgenic potato plants at the mRNA level. / Thesis (M.Sc.)-University of Natal, Pietermaritzburg, 1995.

Ph.D. (Biochemistry) / Genetically modified (GM) crops were first introduced in the 1980s for the production of medicinal products. Since then, areas designated to GM crops have expanded drastically, with the GM crops grown to enhance agricultural productivity, improve agricultural practices, and as a tool to address potential pressures that will be faced by the agricultural sector and to address the issue of food security. Currently, cultivated GM crops include cotton, maize, rapeseed and soybean, carrying agronomic traits such as herbicide tolerance and insect resistance. Following the genetic modification of crops, three possible outcomes can be anticipated: these outcomes include the GM crop produced being equivalent to its untransformed counterpart, the GM crop differing from its untransformed counterpart with several well-defined characteristics, and the GM crop differing from its untransformed counterpart with a multitude of complex characteristics. In cases where the GM crop is equivalent to the untransformed counterpart, no further testing is needed. In instances where several well-defined and characterised differences are found between the GM crop and the untransformed counterpart, safety assessments are performed targeting these differences. The assessments will determine the impact of these unintended and unexpected alterations of the intended enhancement of the GM crops. However, methods currently used to assess GM crops have been found to be lacking, since they only focus on environmental and product-specific risks. Further evidence is essential, as part of GM crop safety assessment, on the molecular characterisation of these crops. This evidence is based on the potential impact of the transformation event, integration of the transgene into the host plant, as well as unintended alterations such as altered gene expression that may occur to the host plant. These events may assist in the further detection of potential dangers of the GM crop. As a result of these highlighted gaps, a project was formulated to study the unintended genomic alterations that may occur during and following the production of a transgenic plant...

Sweet potato (Ipomoea batatas Lam.) is ranked as the seventh most important food crop in the world and its large biomass and nutrient production give it a unique role in famine relief. However, multiple virus infection is the main disease limiting factor in sweet potato production worldwide. The main objective of this research project was to develop a transgenic sweet potato cultivar with broad virus resistance in South Africa (SA). A review of current literature assembled background information pertaining to the origin, distribution and importance of the sweet potato crop; viruses and complexes infecting sweet potato; and the strategies used in sweet potato virus detection and control. A survey to determine the occurrence and distribution of viruses infecting sweet potato (Ipomoea batatas Lam.) was conducted in major sweet potato-growing areas in KwaZulu-Natal (KZN). A total of 84 symptomatic vine samples were collected and graft inoculated onto universal indicator plants, Ipomoea setosa Ker. and Ipomoea nil Lam. Six weeks post inoculation, typical sweet potato virus-like symptoms of chlorotic flecking, severe leaf deformation, stunting, chlorotic mosaic, and distinct interveinal chlorotic patterns were observed on indicator plants. Under the transmission electron microscope (TEM), negatively stained preparations of crude leaf sap and ultra-thin sections from symptomatic grafted I.setosa plants revealed the presence of elongated flexuous particles and pinwheel type inclusions bodies‟ that are characteristic to the cytopathology of Potyviruses. Symptomatic leaf samples from graft-inoculated I. setosa and I. nil were assayed for Sweet potato feathery mottle virus (SPFMV), Sweet potato mild mottle virus (SPMMV), Sweet potato chlorotic stunt virus (SPCSV), Sweet potato chlorotic fleck virus (SPCFV), Sweet potato virus G (SPVG), Sweet potato mild speckling virus (SPMSV), Sweet potato caulimo-like virus (SPCaLV), Sweet potato latent virus (SPLV), Cucumber mosaic virus (CMV), and Sweet potato C-6 virus (C-6) using the nitrocellulose membrane enzyme-linked immunosorbent assay (NCM-ELISA). The majority of leaf samples (52%) tested positive for virus disease and showed the occurrence of SPFMV, SPMMV, SPCSV, SPCFV, SPVG, SPMSV, and SPCaLV. Of these 7 viruses, the most frequently detected were SPFMV (39%), SPVG (30%), followed by SPCSV (13%) and SPMMV (12%). SPCaLV and SPCFV at 10% and SPMSV at 7% were found exclusively in samples collected from one area. SPFMV, SPVG, SPCSV, and SPMMV were identified as the most prevalent viruses infecting sweet potato in KZN. The genetic variability of the three major viruses infecting sweet potato (Ipomoea batatas Lam.) in KZN was determined in this study. A total of 16 virus isolates originating from three different locations (Umbumbulu, Umfume and Umphambanyomi River) in KZN were analyzed. These comprised of 10 isolates of Sweet potato feathery mottle virus (SPFMV), five isolates of Sweet potato virus G (SPVG) and one isolate of Sweet potato chlorotic stunt virus (SPCSV). The phylogenetic relationships of the SPFMV, SPVG and SPCSV isolates from KZN relative to isolates occurring in SA and different parts of the world were assessed. The division of SPFMV into four genetic groups (strains) according to the phylogenetic analysis of coat protein encoding sequences revealed mixed infections of the O (ordinary) and C (common) strains in sweet potato crops from KZN. All SPFMV isolates showed close lineage with isolates from South America, East Asia and Africa. The SPVG isolates showed high relatedness to each other and close lineage with other isolates, especially those from China and Egypt. Analysis of the partial sequence of the Heat shock protein 70 homologue (Hsp70h) gene indicated that the SPCSV isolate from KZN belongs to the West African (WA) strain group of SPCSV and showed close relatedness to an isolate from Argentina. The knowledge of specific viral diversity is essential in developing effective control measures against sweet potato viruses in KZN. Multiple virus infections of Sweet potato feathery mottle virus (SPFMV), Sweet potato chlorotic stunt virus (SPCSV), Sweet potato virus G (SPVG) and Sweet potato mild mottle virus (SPMMV) cause a devastating synergistic disease complex of sweet potato (Ipomoea batatas Lam.) in KZN. In order to address the problem of the multiplicity and synergism of sweet potato viruses in KZN, this study aimed to develop transgenic sweet potato cv. Blesbok with broad virus resistance. An efficient and reproducible plant regeneration protocol for sweet potato (Ipomoea batatas Lam.) cultivar Blesbok was also developed in this study. The effect of different hormone combinations and type of explants on shoot regeneration was evaluated in order to optimize the regeneration protocol. Coat protein (CP) gene segments of SPFMV, SPCSV, SPVG and SPMMV were fused to a silencer DNA, the middle half of the nucleocapsid (N) gene of Tomato spotted wilt virus (TSWV) and used as a chimeric transgene in a sense orientation to induce gene silencing in the transgenic sweet potato. Transformation of apical tips of sweet potato cv. Blesbok was achieved by using Agrobacterium tumefaciens strain LBA4404 harboring a modified binary vector pGA482G carrying the plant expressible neomycin phosphotransferase ll gene (nptll), the bacterial gentamycin-(3)-N-acetyl-transferase gene and the expression cassette. A total of 24 putative transgenic plants were produced from the transformed apical tips via de novo organogenesis and regeneration into plants under 50mg/L kanamycin and 200 mg/L carbenicillin selection. Polymerase chain reaction (PCR) and Southern blot analyses showed that six of the 24 putative transgenic plants were transgenic with two insertion loci and that all plants were derived from the same transgenic event. The six transgenic sweet potato plants were challenged by graft inoculation with SPFMV, SPCSV, SPVG and SPMMV- infected Ipomoea setosa Ker. Although virus presence was detected using NCM-ELISA, all transgenic plants displayed delayed and milder symptoms, of chlorosis and mottle of lower leaves when compared to the untransformed control plants. These results warrant further investigation under field conditions. / Thesis (Ph.D.)-University of KwaZulu-Natal, Pietermaritzburg, 2013.