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Collection of the author's previous publications. Includes bibliographies.

Thesis (MSc)--University of Stellenbosch, 2001. / ENGLISH ABSTRACT: The goals of this project were to (i) determine maximum extractable neutral invertase (NI) activity in the sugarcane culm, (ii) sequence a cDNA encoding for the sugarcane NI (SNI), (iii) determine SNI copy number in the genome, (iv) describe SNI transcript and protein expression patterns throughout the plant, and (v) attempt to determine the contribution of hydrolysis to sucrose accumulation. SNI and sugars were extracted from the developing culm tissues of sugarcane, commercial variety N19. Tissues were divided according to developmental stage (internodes 3, 6 and 9) and anatomical differentiation (enriching for elongating, vascular or storage tissues). The lowest sucrose content was found in the core of the bottom of each of the internodes. The ratio between hexoses and sucrose was highest in the young internodes. In these internodes hexose content was higher in the bottom than the top. There was a significant correlation between sucrose content and NI. Fluxes involved in sucrose synthesis and hydrolysis were investigated. The hexoses glucose and fructose were supplied as a carbon source for tissue discs of young and maturing internodal tissues of sugarcane, varieties N19 and US6656-15. Sucrose content was 10-fold higher in maturing internodes of N19 than US6656-15. Calculated sucrose hydrolysis rates via invertase were higher in maturing internodes of US6656-15 than N19. Taking metabolic compartmentation into account, hydrolysis of sucrose via invertase made a significant contribution to the net turnover of sucrose. Along with this, it would appear that the ability to partition sucrose between the vacuole and cytosol causes a significant difference in sucrose content between varieties. A full-length cDNA for SNI was sequenced. This expressed gene showed significant homology to known NI sequences on both nucleic and amino acid levels. The SNI sequence did not contain the putative invertase catalytic amino acid sequence, suggesting it developed separately from the other classes of invertases. Approximately 1.8 kb of the SNI cDNA was incorporated into a vector suitable for direct bombardment into sugarcane tissue. Southern blot analysis showed the enzyme has a low copy number. SNI transcript expression was observed in all tissues of the sugarcane plant: roots, internodes, leaf roll and leaves. In culm tissues where sucrose content was low and hexose contents were high, SNI transcript and protein levels were high. This suggests that SNI is involved in growth metabolism. / AFRIKAANSE OPSOMMING: Die doel van die projek was om (i) maksimum ekstaheerbare neutrale invertase (NI) aktiwiteit in die suikerriet stingel te bepaal, (ii) die volgorde van 'n eDNA wat vir suikerriet NI (SNI) kodeer te bepaal, (iii) die SNI kopie-getal in die genoom te bepaal, (iv) SNI m- RNA en proteïenuitdrukkingspatrone deur die plant te beskryf, en (v) te poog om die bydrae van hidrolise op sukrose akkumulering te bepaal. SNI en suikers is geëkstraheer uit 'n kommersiële varieteit, N19. Weefsels was volgens ontwikkelingstadiums (internodes 3, 6 en 9) en anatomiese verskille (verryking vir groeiende, vaat- en bergings-weefsels) verdeel. Die laagste sukrose inhoud is in die middel van die onderste helfte van elke internode gevind. Die verhouding van heksoses tot sukrose was die hoogste in die jong internodes. Die inhoud heksoses was hoër in die onderste deel van die internode as die boonste deel. 'n Betekenisvolle korrelasie tussen sukrose inhoud en SNI is gevind. Flukse betrokke by sukrose sintese en hidrolise is ondersoek. Glukose en fruktose is as koolstofbron aan stingelweefsel van twee variëteite (US6656-15 and N19) toegedien. Sukrose-inhoud het tienvoudig tussen volwasse weefsels van die twee variëteite verskil. Hidrolise via invertase was hoër in ouer weefsels van US6656-15 as N19, en het In noemenswaardige bydrae tot sukroseomset gemaak. Die verdeling van sukrose tussen die vakuool en die sitosol kan moontlik 'n groot rol speel in die vermoë van die sel om sukrose te akkumuleer. Die volgorde van 'n volledige SNI eDNA is bepaal. The uitgedrukte geen het, op beide In nukleïen- en aminosuur vlak, betekenisvolle ooreenkoms getoon met ander bekende plant NI volgordes. Die SNI volgorde bevat nie die kenmerkende invertase katalitiese setel nie, wat daarop kan dui dat dit onafhanklik van ander klasse invertases ontwikkel het. Min of meer 1.8 kb van die SNI eDNA is in 'n vektor geskik vir bioliestiese transformering van suikerrietweefsel, geïnkorporeer. Southern klad analise het gewys dat die ensiem 'n lae kopiegetal op geen vlak het. SNI mRNA uitdrukking is waargeneem in elke weefseltipe van die suikerriet plant: wortels, internodes, blaarrol en blare. In stingelweefsels met lae sukrose- en hoë heksose-inhoud, was die vlakke van beide SNI-mRNA en -proteïen hoog. Dit dui daarop dat SNI moontlik betrokke is by groei-metabolisme.

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Thesis (MSc)--University of Stellenbosch, 2006. / Bibliography.

Abstract The effects of dietary sucrose feeding, intragastric sucrose feeding and dietary calcium-deficiency on primary dentinogenesis of the pulp-dentin organ and caries progression were examined in an experimental rat model. The possible role of calcium balance and reduced mineralization of dentin organic matrix as a cause of reduced dentin formation in young, fast growing, rats were also studied. During 3-6 weeks of feeding, immediately after weaning at 3 weeks the urinary calcium, phosphorus, potassium and sodium levels and excretion rates were determined. The areas of dentin formed, the width of the predentin layer, serum mineral and insulin levels, and the areas of dentinal caries lesions were quantified at the end of the experiment. Also the occurrence and progression level of caries lesions were measured. In rat pups, dietary sucrose reduced dentin formation both during the lactation and experimental periods, increased urinary Ca2+ excretions, reduced urinary P, K and Na excretions, and enhanced dental caries occurrence and progression, but it did not affect the width of the predentin layer or the serum mineral and insulin levels. Intragastric sucrose reduced dentin formation and increased Ca2+ excretion, but did not affect the width of the predentin layer, serum mineral and insulin levels in the blood or induce dental caries. Dietary calcium-deficiency reduced dentin formation, increased the width of the predentin layer, caused hypocalcemia, and reduced urinary Ca2+ excretion. These results show that sucrose and calcium-deficiency reduce the rate of primary dentinogenesis through different mechanisms. Calcium imbalance or reduced mineralization of the dentin organic matrix does not explain the reduced dentinogenesis in sucrose fed rats. In conclusion, the present findings indicate that a sucrose load reduces dentinogenesis by impairing the synthesis rate of the dentin organic matrix, but also points out the importance of the local sucrose challenge in initiating dental caries.

Abstract The effects of dietary sucrose and systemic glucocorticoid treatment on the response of the pulpodentinal complex to dental caries were examined in an experimental rat model. The possible role of dentinal caries on dentin formation was also examined. After 5-6 weeks of a dietary and/or medication period, the areas of dentin formation and dentinal caries were quantified in the molars of growing animals. Also the number and severity of caries lesions were estimated. The 43% sucrose diet significantly reduced dentin formation and increased dentinal caries progression. Although glucocorticoid medication alone reduced dentin formation, without dietary sucrose it did not have an effect on caries. In combination of these two, glucocorticoids further increased the progression of dentinal caries, however without significant increase in the number of caries lesions. The cariogenic bacterial inoculation of rats fed a sucrose or control diet increased the progression of dentinal caries. The relationship between cariogenic bacteria and caries was not strong, but there was a stronger relationship between the total amount of dietary sucrose and dentinal caries. In addition to the overall reduction of dentin formation there was no difference in the amount of dentin formed between intact and carious fissures in the sucrose diet group. On the contrary, rats receiving the control diet positively responded to the dentinal caries by increasing dentin formation to prevent pulpal exposure. Whereas the high sucrose diet impaired both the deposition and mineralization of the dentin matrix, glucocorticoids affected matrix formation only. These results indicate that the functional alterations in the pulpo-dentinal complex might contribute to dentinal caries progression in a cariogenic environment, irrespective of the causative mechanism.