D.J. Mycock

In vitro storage of Eucalyptus grandis germplasm under minimal growth conditions.

Slow growth-storage, for up to 10 months, has been achieved for Eucalyptus grandis shoot cultures by either the addition of 10 mg l−1 abscisic acid to the growth medium or by the halving of nutrient supply (half MS) and removal of exogenous plant growth regulators. Reduction of light intensity or the addition of mannitol to the media were less effective in reducing growth rate. Isolated in vitro axillary buds encapsulated in calcium alginate and stored under low temperature and low light intensities survived for up to 3 months without loss in viability. Storage of such encapsulated fresh axillary buds at higher temperature resulted in a loss in viability. These methods have immediate applications to forestry breeding and clonal programs.

Secondary somatic embryogenesis of cassava on picloram supplemented media

The object of this study was to evaluate different strategies for the production of secondary somatic embryos of cassava on picloram-supplemented media. Embryogenically competent calli maintained on double-strength Murashige and Skoog (1962) (MS) medium supplemented with 1 mg l−1 picloram were used as starting material. Secondary embryogenesis from this callus was tested using various basal salt media in either the solid or the liquid state and containing two different concentrations of picloram. Some of the factors effecting the conversion of the embryos into plantlets were also studied. A liquid Schenck and Hildebrand (1972) medium containing 60 g l−1 sucrose and 12 mg l−1 picloram favoured the continual production of a highly embryogenic nodular callus. The normal development of somatic embryos from this tissue was dependant on the use of a picloram-free MS basal salt medium. The embryos were desiccated over a saturated salt solution of K2SO4 (RH 97.5% at 25 °C) and allowed to develop into plantlets on a MS medium containing 0.1 mg l−1 BA. This procedure allowed for the normal elongation of the embryonic hypocotyl and formation of vigorous and viable shoots and roots.

Cryoconservation of South African plant genetic diversity

South Africa has a rich flora which exhibits among the highest species density in the world, distributed across nine biomes that support an impressive diversity of animal life. However, a variety of human actions, invasion by alien species, natural disturbances and climate change collectively impact negatively on the great diversity of both plant and animal species. In situ conservation has long been practised, primarily in nature reserves, complemented by ex situ conservation in national botanic gardens, but in vitro plant conservation is not common. In the context of animal biodiversity conservation, the Wildlife Biological Resource Centre of the National Zoological Gardens utilises cryobanking as one of its major focuses and is now poised to expand as the repository for the cryoconservation of plant germplasm, particularly for indigenous recalcitrant-seeded and poor-seeding species. However, there are particular problems associated with successful germplasm cryostorage of such tropical and subtropical plants. As we see the science and application of cryobiology and cryoconservation as cross-cutting and transdisciplinary, we have entrained formal networking among scientists offering a range of specialisations aimed at a deeper understanding of common problems and practical outcomes to facilitate both plant and animal biobanking. The endeavours are aimed at elucidating the basis of both successes and failures in our efforts to attain optimal outcomes. With focus on best practices, standard operating procedures, validation and risk management for cryopreserved and cold-stored plant and animal material, our ultimate aim is to facilitate restoration by the safe reintroduction of indigenous species.

Development of Cassava (Manihot esculenta Crantz.) somatic embryos during culture with abscisic acid and activated charcoal

Summary Secondary cassava somatic embryos were cultured on a sequence of developmental media containing either activated charcoal or abscisic acid. Medium supplementation with activated charcoal had a positive effect on both differentiation and subsequent germination. However, sustained exposure to activated charcoal supplemented media beyond the seventh day of differentiation did not appear to be beneficial. Somatic embryos were desiccated over a saturated salt solution of K 2 SO 4 for 10 days and allowed to develop into plantlets on a MS medium containing 0.4 μmol/L BAP. Embryo germination was significantly affected by the culture medium used prior to the desiccation treatment, with culture on a medium supplemented with ABA at a level of 0.1 μmol/L prior to desiccation significantly enhancing germination. The number of normal germinants produced as judged by morphological criteria were not however, dependant on ABA supplementation of the culture medium prior to desiccation. Culture on a separate medium for the differentiation of embryos, different to the medium used for the maturation of somatic embryos is indicated.

Applications of in vitro methods to Eucalyptus germplasm conservation

SYNOPSIS For forestry species that require large tracts of land for conservation stands, in vitro storage methods such as minimal growth cultures or cryopreservation can provide a cost- and space-effective ex situ conservation approach. As protocols for the micropropagation of Eucalyptus spp. are available, collections can be rapidly retrieved from in vitro storage, multiplied, and be ready for distribution throughout the year. However, one of the major disadvantages of this approach is that the methods for storage have to be developed for each species (and sometimes clones) and this is initially time consuming and expensive.

Cryopreservation of Somatic Embryoids of Phoenix dactylifera

Late globular/early torpedo stage date palm embryoids can continue normal growth and development after cryopreservation provided they are pre—treated with a cryoprotectant mixture of glycerol and sucrose and then dried to water contents in the range of 0.4–0.7g.g-1. The embryoids were frozen by direct immersion in liquid nitrogen. Although further drying allows for 100% recovery, growth is in the form of unorganized callus. Tissues frozen at extremely rapid rates (by immersion in liquid freon) retain cytoskeletal structures, whereas material frozen at slower rates (in liquid nitrogen) appear to lose this subcellular system. The slow recovery rate of material frozen in liquid nitrogen could, in part, be due to reconstitution of this subcellular matrix.

General applicability of in vitro storage technology to the conservation and maintenance of plant germplasm

The broad applicability of in vitro storage technology to the preservation of germplasm for agriculture, horticulture, forestry, biotechnologically based industries and the conservation of plants that are endangered forms the basis of this contribution. In order for this technology to be implemented it is necessary to have efficient and reliable micropropagatory procedures for the species under question. The development of both sets of procedures are discussed for cassava, Eucalyptus and two endangered indigenous Haworthia species. In each case storage is required for different reasons and the versatility of the in vitro storage technology in satisfying the requirements is highlighted

The salt glands of Tamarix usneoides E. Mey. ex Bunge (South African Salt Cedar).

ABSTRACT Tamarix usneoides is a halophyte tree endemic to south-western Africa. This species is known to excrete a range of ions from specialized glandular structures on its leaves. To understand the mechanisms involved in the transport, sequestration and excretion of ions by the glands, a study was performed on salt gland distribution and ultrastructure. The glands are vesiculated trichomes, comprised of eight cells viz. two basal collecting cells and six excretory cells, partially bounded by a secondary cell wall that could serve as an impermeable barrier, forcing excess ions to move from the apoplast of the surrounding tissue into the cytoplasm of the basal excretory cells. It was hypothesized that the ions are moved across the excretory cells in endocytotic vesicles that fuse with the plasmalemma or form junctional complexes, allowing ion movement from one excretory cell to the next. In the apical cell, the vesicles fuse with the plasmalemma, releasing the ions into the network of cell wall ingrowths which channel the ions to the outside surface of the cell. This study shows that there are distinct structural adaptations for the processing of ions for excretion, although the mechanism by which ions enter the cells still needs to be determined.

Cryostorage of Pea (Pisum sativum L.)

The common pea (Pisum sativum L.) is probably one of the best known legumes used as a fresh vegetable (Langer and Hill 1982). It is not known in the wild state, but is very closely related to P. arvense, which occurs in Georgia, Russia (Purseglove 1968; Janick et al. 1981). Pea appears to have been first cultivated about 7000 B.C. in the south-western parts of Asia, the same general region from which wheat originated (Janick et al. 1981). It was spread west to the Greeks, who passed it on to the Romans and thence to the Germanic races of Europe. Prior to European global migration, pea cultivation had also spread east to India and China and south into central Africa (Purseglove 1968; Janick et al. 1981). During the 18th century, Gregor Mendel, in his classic work on the genetics of pea, recognized 32 different types. Today, about 10 000 cultivars have been recorded, most from temperate areas of the world where the crop is grown best (Gantotti and Kartha 1986).