In South Africa, Eucalyptus trees are used for many processed wood products (e.g. paper) and in the mining industry. Priorities in Eucalyptus breeding programmes include selection of varieties that are fast growers, insect and disease resistant, have appropriate wood characteristics and can grow in a wide variety of environmental conditions. Cold-tolerant cultivars of E. saligna and of E. grandis have been bred and selected in Australia and South Africa, respectively for use in cold regions of Natal Midlands and North Eastern Cape. However, the production of large numbers of such cultivars for planting out in a commercial scale is being impaired by slow growth rate, low regeneration time and poor rooting ability of cuttings from these trees. Consequently, methods of in vitro propagation of cold-tolerant clones were investigated. Axillary buds were induced and subjected to a variety of multiplication, elongation and rooting media. The optimised protocol for the production of shoots from axillary buds was: bud induction medium comprising of MS supplemented with 20 grl sucrose and 10 grl agar for 1-2 weeks, multiplication medium comprising of MS supplemented with 0.1 mgrl biotin, 0.1 mgrl calcium pantothenate, 0.2 mgrl benzyladenine phosphate, 20 grl sucrose and 3.5 grl Gelrite for 4 weeks, elongation medium for 4-6 weeks comprising of MS medium supplemented with 0.1 mgrl biotin, 0.1 mgrl calcium pantothenate, 0.35 mgr' NAA, 0.1 mgr' kinetin, 0.1 mgrl IBA, 20 grl sucrose and 3.5 gr1 Gelrite. Production of plantlets via somatic embryogenesis was also investigated but hampered because of high rates of contamination as pieces of mature leaves were used as exp1ants. Ongoing breeding programmes are aimed at obtaining hybrids of Eucalyptus that are cold tolerant. The hybrid progeny then need to be screened for cold-tolerance. However, a major problem in the selection of cold-tolerant clones is that diagnosis can only be undertaken by assessing the field performance of the genotypes under various environmental conditions. In this regard, a protocol for 1D gel electrophoresis was developed for Eucalyptus species with the view to use it for the detection of cold-tolerant stress proteins. Leaf material from both non-cold tolerant and cold-tolerant clones was used. Well-resolved gels that focused on the comparison' between protein profiles of cold-susceptible and cold-tolerant clones before and after period of cold stress were obtained. The findings of this study showed that two polypeptides, one in the lower molecular region of 14.3-20.1kD and another of a higher molecular weight in the region of 116.4-170 kD were observed after cold acclimation. These changes in polypeptide profiles were observed in cold-tolerant E. grandis x nitens (GN1) and E. saligna (AS 184, AS 196 and TS 15) but not in a non-cold tolerant species E. grandis (TAG 731). These polypeptides may have an important role in the cellular adaptation to cold temperatures. It is suggested that this method may be used as a diagnostic tool for screening cold tolerance on Eucalyptus cultivars.

Thesis (M.Sc.)-University of Natal, Durban, 1996.