The two species investigated in this study were Theobroma cacao and Barringtonia racemosa. Theobroma cacao has worldwide economic importance, as cocoa (the main ingredient in chocolate) is produced from the seeds of this tree; while B. racemosa has several applications in herbal medicine. The seeds of both T. cacao and B. racemosa are highly recalcitrant and therefore not amenable to storage for any significant periods. The long-term conservation of the germplasm of these species may only be feasible via cryopreservation. The aims of the present study were to: 1) optimize in vitro regeneration protocols for different types of explants that have the potential to be cryopreserved while maintaining the genetic integrity of these two species; and 2) develop cryopreservation protocols for selected explants. For T. cacao, protocols were established for bud-break and multiplication for both in vitro - and greenhouse-derived nodal explants, as well as a rooting medium for shoots derived from axillary buds. Parameters investigated towards the cryopreservation of axillary shoots, from greenhouse nodal segments, and nodal segments from in vitro plantlets, included the size of the explant and pre-treatments for cryopreservation. Nodal segments (6 - 7 mm) and axillary shoots (2 - 4 mm) needed to be soaked in 0.5% (w/v) ascorbic acid for 10 min to minimise phenolic production and subsequent tissue death, and surface-sterilized by soaking in 1% Ca(OCl)2 solution for 5 min to reduce microbial contamination. Subsequent cryopreservation attempts involved only in vitro nodal segments because of the lack of success in achieving elongation of excised axillary buds. Vitrification and slow freezing methods, with or without the application of cryoprotectants, did not achieve successful cryopreservation. Attempts to establish a protocol for producing somatic embryos, as an alternate to axillary shoots and in vitro nodal segments, resulted in the production of globular embryogenic callus for both leaf and cotyledon explants. Cryopreservation of these explants was not investigated in the scope of this study. The study on B. racemosa focused on the development of a somatic embryogenesis protocol. Segments of embryonic axes produced globular-stage embryos when placed on MS medium supplemented with 30 g 1-1 sucrose, 1.0 g 1-1 casein hydrolysate, 2.0 mg 1-1 2,4-D, 0.1 mg 1-1 BAP and 8.0 g 1-1 agar. Various strategies were employed to obtain embryo germination, which included 1) different time intervals on callus initiation medium; 2) the use of different auxins (IAA, NAA and 2,4-D) in combination with the cytokinins BAP and kinetin; 3) desiccation and 4) cold treatments. Although somatic embryo germination was not achieved, globular embryos proceeded with development to the cotyledonary stage when cold-treated for 8 h at 4°C. This study provides some fundamental bases for further investigation towards achieving long-term conservation for both T. cacao and B. racemosa. However, the use of meristems as explants for cryopreservation is suggested to be the way forward for the cryopreservation of both species. / Thesis (M.Sc.)-University of KwaZulu-Natal, 2008.

Seeds remain the most convenient and successful way for storing the genetic diversity of plant species and for producing new plants routinely for agriculture and horticulture. The importance of seed storage and the ability to predict seed longevity must therefore not be underestimated. To be successful, storage conditions must maintain seed vigour and viability and ensure that normal seedlings are subsequently established under field conditions. Seed quality is best retained when deteriorative events are minimised, which is achieved by storage of low moisture-content seeds under cool to cold, or even sub-zero, temperatures. Such conditions are employed for 'orthodox' seeds, which are desiccation tolerant and able to survive at sub-zero temperatures in the dehydrated state for extended periods. It is seeds referred to as 'recalcitrant' that cannot be dehydrated and often not stored at low temperatures because they are desiccation sensitive and may not tolerate chilling. According to almost anecdotal records chilling temperatures for such seeds are those below 15°C down to 0°C, depending on the species. The limited storage lifespan of recalcitrant seeds presents a problem even for short-term storage, and as most research on chilling sensitivity has been conducted on vegetative tissue, relatively little data exist for seeds, especially recalcitrant types. The purpose of this study was to gain an understanding of the chilling response of recalcitrant seeds, as reduced temperature could have the potential to extend, rather than curtail, storage lifespan, depending on the species. Selected physiological, biochemical and ultrastructural responses of recalcitrant seeds of Avicennia marina and Ekebergia capensis were characterised. Seeds of the two species were stored at 25, 16 and 6°C. Germination, water content (determined gravimetrically), respiration (measured as CO2 production) and leachate conductivity (tissue electrolyte leakage over time) were assessed at regular intervals. Chilling response at the subcellular level was examined using transmission electron microscopy (TEM). Changes in sugar metabolism and activities of the antioxidant enzymes superoxide dismutase (SOD), catalase (CAT) and glutathione reductase (GR) were assessed for A. marina seeds, which were severely affected by the chilling temperature of 6°C, losing viability after 1 week. In contrast, the seeds of E. capensis retained viability after 12 weeks of storage at 6°C, indicating the marked difference in chilling response between seeds of the two recalcitrant species, despite their common tropical provenance. However, when E. capensis seeds were stored at 3°C viability decreased significantly after 8 weeks, thus indicating how critically temperature must be controlled if such conditions are to be profitably employed. Ultrastructural studies revealed that in both E. capensis and A. marina seeds vacuole formation was initiated more rapidly at lower temperatures than at higher temperatures, indicating that this was a response specific to the chilling stress imposed. Once again, 'lower temperatures' differed relative to the species concerned. In the E. capensis seeds, nucleolar morphology was affected and the extent of chromatin patches in the nuclei increased as the storage temperature was reduced. Other ultrastructural findings could not be linked specifically to the chilling stress imposed on the E. capensis and A. marina seeds. Activity of the antioxidant enzymes SOD and GR was detected in the A. marina seeds. No measurable CAT activity was detected. Glutathione reductase activity increased in response to chilling stress, the rate of the increase depending upon the severity of the chilling stress imposed. Other than when the A. marina seeds were placed directly at 6°C, there were no notable increases in SOD activity. Interestingly, SOD and GR activity was not the same in the axes as in the cotyledons. Superoxide dismutase activity was found to be higher in the axes and GR activity higher in the cotyledons. It would have been beneficial to determine the extent of antioxidant enzyme activity in the E. capensis seeds as well if this had been possible. Generally, chilling of recalcitrant seeds seems to evoke a response similar to that of dehydration below a critical water content. This could lead to the conclusion that recalcitrant seeds do not possess the genetic ability to cope with dehydration or chilling stress, if it were not for the existence of recalcitrant seed species that are more chilling tolerant. / Thesis (M.Sc.)-University of Natal, Durban, 2002.

The consequences of toxigenic fungi associated with stored seed have stimulated these investigations aimed at developing treatments to minimise this mycoflora, without significantly reducing seed quality or viability. The effects of immersion in water at 55, 57 and 60 QC for durations of 5 to 60 min were assessed for maize (Zea mays L.) seed in terms of fungal status, water uptake, electrolyte leakage, germination and seedling establishment. These assessments were conducted immediately after treatment, after re-dehydration for 2 days in an ambient air stream, and following a 1 month storage period under either cold (4 QC) or ambient (25 QC) conditions (33% and 91% RH, respectively). In all cases, the results are compared with those of control seeds and seeds pre-imbibed for 4 h at ambient temperature. The level of internal contamination, represented almost entirely by Fusarium moniliforme Sheldon, declined significantly when assessed immediately after treatment, the efficacy of which increased with increasing temperature and duration of treatment. Seeds immersed in water at 55 QC for a duration of 15 min exhibited an 85% reduction in infection levels, when compared with those of the control, while those treated at 57 and 60 QC (same duration) were uninfected. Immersing seeds in hot water, however, resulted in a lag in germination rate and drop in germination totality, the degree of which was enhanced by increasing duration and temperature of treatment, suggesting the status of the manipulation to be an accelerated ageing treatment. The electrolyte leakage studies indicated that the reduced germination performance of these seeds was not due to plasmalemma disorganisation. These deleterious effects, however, were counter-balanced as seeds treated at 55, 57 and 60 QC for durations up to 60, 30 and 10 min, respectively, produced plants of superior quality than those of the control, which is ascribed to the reduction of systemically transmitted pathogens. The efficacy of the hot water treatment in reducing the levels of seed infection and improving seedling quality was enhanced by subsequent re-dehydration. The reduction in seed-associated mycoflora was maintained following storage for 1 month at both 4 QC (33% RH) and 25 QC (91% RH). However, both seed and seedling quality were adversely affected following storage even under cold, dry conditions, which may be a consequence of the pre-treatment history of the seeds, which had been cold-stored for two years prior to the experiments. Applied as a pre-sowing treatment, therefore, hot water treatment shows promise for producing a crop of superior quality, less prone to fusarial pathogenesis. This treatment may be of particular importance to Third-World subsistence communities. / Thesis (M.Sc.)-University of Natal, 1995.

Bibliography: leaves 155-182. / xiv, 182, [10] leaves, [11] leaves of plates : ill. (some col.) ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Most agricultural crop seeds are 'orthodox' ie. desiccation is a necessary feature of their complete life cycle. Low moisture content lengthens the storage life of orthodox seeds. Recalcitrant seeds, which do not tolerate low moisture content, lose viability in dry storage. The thesis studies the role of sugars in desiccation tolerance in developing seeds and investigates other mechanisms which might be involved in desiccation tolerance and desiccation sensitivity. / Thesis (Ph.D.)--University of Adelaide, Dept. of Plant Science, 1996?

Stage of maturity at harvest and relative seed size can affect seed vigor. Greenhouse and field studies were conducted to determine the effect of seed maturity on seed vigor, storability, and subsequent plant growth of four cultivars of snap bean (Topcrop, Provider Black, Provider White, and Cherokee Wax). Seeds harvested between physiological maturity (PM) and harvest maturity (HM) showed the highest seed vigor and storability. At PM, seed moisture content was about 55%. A drop in seed fresh weight and a pod color change from green to yellow appear to signal the stage of PM. Delaying harvest past HM reduced seed vigor. The three cultivars with colored seed coats showed higher seed vigor than Provider White. Climatic and weathering effects on maturity, vigor, and yield of Topcrop and Cherokee Wax seed were evaluated at three locations (L-I, L-2, and L-3) in Sri Lanka having different agro-climatic conditions. Seeds that developed and matured under cooler conditions (L-l) produced higher yields and had larger size, better color, and higher vigor. Seeds developed and matured faster under warmer conditions (L-3); however, seed vigor and yields were lower and seed vigor dropped prior to HM. Cherokee Wax was the most tolerant to field weathering. Seeds of Topcrop and Cherokee Wax were separated by weight into three seed sizes (small, medium, and large) and evaluated for crop performance in two plantings (12/90 and 3/91). Field emergence was higher from larger seeds in the second planting, where the soil conditions were more stressful. Seedlings and plants at the flowering stage were larger and pod yields higher from larger seeds. The cultivar x seed size interaction was significant for pod yield per plant. Topcrop showed no differences, while with Cherokee Wax, pod yield per plant increased with increase in seed size during both plantings. Seeds produced from small seeds were similar or higher in vigor, indicating that small seeds could be used for seed production purposes. The data from these experiments indicate that vigor in snap bean seeds can be optimized by harvesting at an early stage after PM and by grading to remove small seeds. / Ph. D.

Call number: LD2668 .T4 1985 M62 / Master of Science

Call number: LD2668 .T4 1985 M63 / Master of Science

Graduation date: 1994

Biochemical and ultrastructural changes associated with chilling injury (CI) in soybean seeds imbibed at 5°C and 25°C were investigated. Soybean seed germination appeared to be affected by chilling temperatures and initial seed moisture content. Seeds with higher moisture contents exhibited 85% germination, while low moisture content seeds had a 32% germination. Leakage rates were greater in chilled seeds, indicating that membrane integrity in the tissues was impaired at chilling. The low rates of potassium ion leakage between 6 and 24 hours of imbibition compared to the high peroxide levels observed during this period led to the suggestion that lipid peroxidation was a better marker of CI than leakage. Transient changes in lipid hydroperoxide levels were observed in chilled and non-chilled seeds and axes. However, in axes, the increase in lipid hydroperoxides after 12 hours of imbibition at chilling temperatures was associated with an 18% decline in linoleic acid levels of total lipid fraction. Similarly, a 10% decline was observed in the polar lipid fraction. These results suggest that the capacity of seeds to control lipid peroxidation may be an important component in CI and that a consequence of peroxidation is likely to be a loss of fatty acid unsaturation. Sugar levels were not affected by chilling and non-chilling temperatures and no relationship could be established with CI. Antioxidant defense enzymes (catalase and superoxide dismutase) were expressed at chilling and non-chilling temperatures and increases were observed after 24 hours of imbibition which showed an apparent correlation with increases to lipid hydroperoxide levels. Enzyme levels decreased after 48 hours of imbibition at a time which coincided with the decline observed in the peroxide levels. Overall, no marked differences were observed in chilled and non-chilled cells at the ultrastructural level, except that vacuolar reserve mobilization was markedly impeded. / Thesis (M.Sc.)-University of Natal, Pietermaritzburg, 1996.

No description available.