Doctoral Degrees (Botany)
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Browsing Doctoral Degrees (Botany) by Author "Bornman, Chris H."
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Item Abscisic acid and other hormonal effects on growth in Spirodela.(1969) Van Staden, Johannes.; Bornman, Chris H.The effects of abscisic acid In particular, as well well as gibberellic acid and the cytoklnlns, 6-benzyladenine, kinetin, and 6-dlmethylal lalylamlnopurine, on the growth of Spirodela oligorrhiza were investigated. Abscisic acid effectively arrested growth permanently at concentrations down to 10ĀÆĀ¹ mg/I. Normal growth tended to be resumed at concentrations of 10ĀÆĀ² and 10ĀÆĀ³ mg/l between nine and twelve days after treatment. A concentration of 10[-8] mg/l, however, resulted in a significant increase in dry weight at both eight, nine and twelve days after introduction into the culture medium. It is suggested that the resumption of growth twelve days after treatment at those concentrations which inhibit growth up to nine days, was due to a possible progressive inactivation of abscisic acid resulting in a lowering of its concentration to a level that is promotive. It was furthermore found that the growth response of Spirodela in terms of dry weight production over a period of eight days is proportional to the log[10] concentration of abscisic acid. It is suggested that this curve can be used as a relatively reliable and easily performed bioassay to detect amounts of abscisic acid as low as 10[-5] Ī¼g. The assay is more reliable over the range 0.01 to 10,000 Ī¼g and appears not to be affected by gibberellin, benzyladenine and kinetin. The inhibitory effect of abscisic acid on growth in Spirodela was shown to be reversed by benzyladenine, kinetin and dimethylallalylaminopurine, although they were not equally effective in doing so. Benzyladenine at 1.0 mg/l was the most effective In overcoming growth inhibition by abscisic acid. Gibberellic acid, however, proved ineffective in reversing the inhibitory effect of abscisic acid on Spirodela oligorrhiza. The apparent Increases in growth obtained in some cases may have resulted more directly from gibberellic acid stimulation than from the Interaction of gibberel lie acid with abscisic acid.Item Studies on the primary phloem of Gomphocarpus physocarpus.(1974) Botha, Christiaan Edward Johannes.; Bornman, Chris H.Penetration of the stems and leaves of Gomphocnrpus physocarpus by the aphid, Aphis nerii was studied with light and phase microscopes. Penetration of the epidermis and ground tissue was largely intercellular, that of the phloem tissues partly intercellular and in part intracellular. Of 75 pairs of stylet tips encountered in presumably functional sieve tubes 73 were lodged in sieve tubes of the internal and adaxial phloem. Light and electron microscope investigation of the ontogeny and differentiation of the primary phloem revealed that the internal and adaxial phloem attains a more advanced stage of maturity earlier than the corresponding external and abaxial strands. Furthermore, the protoadaxial phloem sieve tubes remain functional throughout the primary stages of growth, whereas the corresponding abaxial phloem sieve plates are associated with massive deposits of definitive callose during the latter stages of primary growth. The internal and adaxial phloem was found to be capable of independent assimilate translocation in plants which had the external phloem experimentally ringbarked. Translocation in the internal and adaxial phloem of such plants was unaffected by ringbarking. The results are interpreted as indicating that a greater proportion of assimilates is translocated in the internal and adaxial phloem sieve tubes than in the external and abaxial sieve tubes. It is concluded that this governs A. nerii's preference for the internal and adaxial phloem as a prime feeding site.Item Tissue culture studies on citrus and Welwitschia.(1972) Button, James.; Bornman, Chris H.Part I. IN VITRO CULTURE OF CITRUS EMBRYOS AND NUCELLAR ISOLATES Zygotic embryos of the Ellendale mandarin, a monoembryonic variety of citrus, were cultured on modified basal media of Murashige and Skoog (BM[1]), and White (BM[2]) , supplemented with various growth regulators and nutrient additives. The growth of immature embryos was greatly enhanced by the addition of 400 mg/l casein hydrolysate (CH) to the basal media. Coconut milk (CM) and malt extract (ME) enhanced growth to a lesser extent, while the addition of indoleacetic acid (IAA) and kinetin (KIN) at the concentrations used, was in no way beneficial. Nucellar isolates excised from abortive and normal Ellendale mandarin ovules eight to 20 weeks after anthesis, were cultured on BM[I] and BM[2] in the presence of various concentrations and combinations of IAA, indolebutyric acid, naph~haleneacetic acid (NAA), 2,4-dichlorophenoxyacetic acid (2,4-D), KIN, CM, benzyl adenine, 6-dimethylallylamino purine, yeast extract (YE), ME, CR, adenine (AD), adenine sulphate (AS), ascorbic acid (AA), and benzylthiazole-2-oxyacetic acid. Some of the isolates which remained alive for four months did develop callus but n~ differentiation of embryoids or other structures occurred. Unfertilized ovules from 8-12-week-old Washington Navel orange fruits provided nucellar isolates which were cultured on media similar to those upon which mandarin nucelli were unsuccessfully cultured. In the case of Navel orange nucelli however, BM[1] + 400 mg/l filter-sterilized ME, and BMl + 40 mg/l AD yielded numerous pseudobulbils which later developed embryoids. Adenine (10 mg/l) was more effective than 20 mg/l which in turn was more effective than 30 mg/l. Adenine was more effective than its equivalent amount supplemented in the sulphate form except at 10 mg/l where the two forms were equally effective. Zeatin (ZE) at 0,2 mg/l did induce some pseudobulbils and embryoids, but all these treatments were less effective than 400 mg/l ME. When transferred to BM[1] + GA[3] (1 mg/l) , embryoids developed roots and later, shoots. It was necessary to remove plantlets from the GA[3]-supplemented medium shortly after the first foliage leaves developed in order to prevent the development of weak, spindly plants . Plantlets were transferred from BM[1 ]+ GA[3] to BM[1] only, and then after careful conditioning they were planted out in soil. This appears to be the first successful attempt at inducing adventive embryogenesis 1n the nucellus of unpollinated, unfertilized citrus ovules in vitro. Part II. EMBRYO AND FREE-CELL CULTURE OF WELWITSCHIA MIRABILIS Welwitschia embryos, cultured on BMI supplemented with CR, and low levels of IAA and KIN, germinated and developed leaves but not roots. Embryos cultured on BM[I] with 5,0 and 10,0 mg/l NAA produced an abundance of friable callus from the hypocotyl root axis. This callus was used for starting suspension cultures aimed at inducing vegetative embryogenesis. A number of nutritional additives and hormones were used alone and in combination at various concentrations. Cells of numerous shapes and sizes were observed but no organogenesis was apparent in either suspension cultures or in cell colonies plated out on semi-solid agar media. A closer study of cell aggregates formed in suspensions supplemented with CM + 2,4-D revealed that internal division occurred in approximately 40 per cent of the larger cells. It is suggested that this internal division may constitute the first step in embryogenesis of Welwitsahia cells in suspension culture. It is also tempting to speculate that this process, which has been reported by other researchers, is the first step 1n embryogenesis of free cells in general. Although this attempt at inducing adventive embryogenesis in cell cultures of Welwitschia was unsuccessful, some encouraging results were obtained on potentially suitable media and possible initial stages in the organization of embryoids.Item Ultrastructure of the germinating Welwitschia mirabilis seed.(1975) Butler, Valerie.; Bornman, Chris H.The structure and chemical composition of quiescent Welwitschia mirabilis (Hooker fil.) seeds and resultant changes occurring during the first 7 days of germination were studied. Research was focussed on the megagametophyte and its interrelationship with the non-vascularised outgrowth produced by the embryo. This outgrowth was given the name "feeder" by Bower in 1881 who postulated that it functions as an absorptive organ. However the possibility existed that it merely fulfilled the mechanical role of anchorage. Following hydration activation of embryonic collar cells precedes that of gametophyte cells whose rate of activation is governed by relative distance from the embryo. This sequence of activation is suggestive of a stimulatory factor diffusing from the embryo into the gametophyte. Starch, protein and lipid reserves in the collar and developing feeder are consumed within 36 to 48 h. As a consequence the rapidly developing seedling is probably largely dependent on nutrient material in the gametophyte until the plumule emerges, after approximately 5 to 6 days germination. Ventral feeder cells in contact with gametophyte tissue apparently act as transfer cells, developing numerous small wall projections invested with plasmalemma which result in a much greater absorptive surface area. The large numbers of mitochondria occurring in these cells might suggest active uptake of nutrients. At the 3- to 4-day-stage the feeder and gametophyte adhere firmly. While this adherance probably facilitates translocation of nutrients it could also have the secondary function of anchoring the feeder in the gametophyte, thus providing the emerging plumule with a firm base. The apparent root cap origin of ventral feeder cells might explain the positive geotropism of the feeder, as recent work inter alia by Wilkins and Wain (1975) has shown that root cap cells may be geoperceptive. Cytochemical methods used at the light and electron microscope level suggest that reserve material within protein bodies of the embryo and gametophyte might exist as a proteincarbohydrate complex and that globoid origin might be cytoplasmic. The immediate digestion of protein body reserves in the embryo and gametophyte interface zone argues the presence of pre-existing hydrolytic enzymes laid down within the protein bodies prior to quiescence. However the enzymes responsible for reserve breakdown in deep gametophyte tissue seem to be synthesised de nova. Protein hydrolysis precedes lipid digestion which possibly indicates that some of the resulting free amino acids might be used in the de nova synthesis of lipases. Lipid bodies, microbodies, mitochondria and amyloplasts encircled with ER seem to form a complex. Fatty acids resulting from lipase action in the lipid bodies (Ching 1968) are probably converted by microbodies (glyoxysomes) to succunate (Breidenbach and Beevers 1967) which is converted to sucrose by the action of mitochondria (Cooper and Beevers 1969a, b). Excess sucrose is probably converted to starch and stored in the amyloplasts. In 5 days the mean dry mass of the gametophyte decreases by approximately 47% during which time the total amount of lipid decreases by 76.5% and protein by 14%. Although some of the hydrolysed fatty acids and amino acids are no doubt utilised in the gametophyte it is suggested that the majority of fatty acids are probably converted to sugarsĀ· which, together with free amino acids (and possibly simple peptides) are transported to and absorbed by the embryo via the feeder whence they are utilised for seedling growth.