Development of micropropagation protocols for selected indigenous plant species..

Abstract

The herbal medicine trade is thriving in KwaZulu Natal with an ever-increasing number of people harvesting and trading in indigenous plants, especially those species with medicinal and/or magical properties. The number of plants harvested has increased whereas the size of the plants collected has decreased, resulting in low recruitment into wild populations. As a result of these two factors, species diversity has decreased. To this end, the aim of these investigations was to establish micropropagation protocols for the selected species i.e. Bowiea volubilis, Haworthia_ limifolia and Cryptocarya latifolia. In addition, hardening-off protocols were also developed. The bulbous plant, Bowiea volubilis, was propagated via organogenesis using the inflorescence stem. Bulblet formation occurred directly without an intervening callus phase. Bulblets were produced on explants on Linsmaier and Skoog (1965) (LS) medium containing 30 g.r' sucrose and either I mg.r' BAP and I mg.r' 2,4-D or 1 mg.r' BAP and 1 mg.r' NAA. Shoots and roots were induced upon transfer to the basal medium devoid of plant growth regulators. Regenerated plantlets were successfully hardened-off. Haworthia limifolia, a succulent, was propagated via direct somatic embryogenesis using leaf material. Embryo formation was induced on a modified Murashige and Skoog (1962) (MS) medium containing 20 g.r' sucrose and 1 - 5 mg.r' 2,4-D. secondary embryogenesis occurred when the explants were transferred to the basal medium supplemented with activated charcoal and devoid of growth hormones. Healthy plantlets, produced from secondary embryos, were transferred to pots and acclimatised to greenhouse conditions. A large proportion of the plantlets regenerated were vitrified and as a result, this problem was addressed by changing the medium composition or culture environment. Silica gel, when placed in the culture vessel, was the best treatment for reversal of the vitrified condition. The establishment of leaf and nodal segment cultures of Cryptocarya latifolia required extensive investigation of sterilants to reduce fungal contamination. Several fungicides were tested and a successful sterilisation protocol was established. A number of media were tested for the induction of dormant axillary buds and multiplication of shoots. The best medium for both bud induction and proliferation was MS medium containing 30 g.r1 sucrose and 1 mg.r1 BAP and 0.01 mg.r1 NAA. Callus cultures were established on MS medium containing 30 g.r1 sucrose and 3 mg.rl 2,4-D. These calli, however, were non-embryogenic. Application of the established protocols and future research strategies are discussed.