History

Although shoot culture has proved to be a widely applicable method of micropropagation, the appreciation of its potential value developed only slowly, and utilisation largely depended on improvements in tissue culture technology. Robbins (1922) seems to have been the first person to have successfully cultured excised shoot tips on a medium containing sugar. Tip explants of between 1.75 and 3.75 mm were taken from pea, corn and cotton, and placed in a liquid medium. For some reason the cultures were maintained in the dark where they only produced shoots with small chlorotic leaves and numerous roots. Although it is tempting to suppose that the potential of shoot culture for plant propagation might have been appreciated at a much earlier date had the cultures been transferred to the light, the rapid rate of shoot multiplication achieved in modern use of this technique depends on later developments in plant science.

Only very slow progress in shoot culture was made during the next 20 years. As part of his pioneering work on plant tissue culture, White (1933) experimented with small meristem tips (0.1 mm or less) of chickweed (Stellaria media), but they were only maintained in hanging drops of nutrient solution. Leaf or flower primordia were observed to develop over a six-week period. Shoot culture of a kind was also carried out by La Rue (1936). His explants largely consisted of the basal and upper halves of seed embryos. Nevertheless, the apical plumular meristems of several plants were grown to produce entire plants. Whole plants were also obtained from axillary buds of the aquatic plant Radicula aquatica.

Significant shoot growth from vegetative shoot tip explants was first achieved by Loo, and reported in 1945 and 1946 a, b. Asparagus shoot tips 5-10 mm in length were supported on glass wool over a liquid medium and later grown on a solidified substrate.

Loo (1945, 1946a, 1946b) made several significant observations showing that:

• growth depended on sucrose concentration, higher levels being necessary in the dark than in the light;

• explants, instead of being supported, could be grown satisfactorily on 0.5% agar;

• in vitro shoot growth could apparently be continued indefinitely (35 transfers were made over 22 months);

• shoot tip culture afforded a way to propagate plant material (clones were established from several excised shoot apices).

This work failed to progress further because no roots were formed on the Asparagus shoots in culture. Honours for establishing the principles of modern shoot culture must therefore be shared between Loo and Ball. Ball (1946) was the first person to produce rooted shoots from cultured shoot apices. His explants consisted of an apical meristem and 2-3 leaf primordia. There was no shoot multiplication but plantlets of nasturtium (Tropaeolum majus) and white lupine (Lupinus alba) were transferred to soil and grown successfully.

During several subsequent years, shoot apex (or meristem tip) culture was of interest only to plant pathologists who recognised its value for producing virus-tested plants. It was during studies of this kind that Morel made the significant discovery of protocorm formation from Cymbidium orchid shoot tips. Although they may be started from the same explants, cultures giving rise to protocorms are not typical shoot cultures (see later in this chapter).

The two major developments which made shoot culture feasible were the development of improved media for plant tissue culture (Murashige and Skoog, 1962) and the discovery of the cytokinins as a class of plant growth regulators (Miller, 1961b; Skoog et al., 1965), with an ability to release lateral buds from dormancy (Wickson and Thimann, 1958; Sachs and Thimann, 1964). These developments were not immediately applied to shoot culture, and some years elapsed before it was appreciated that multiple shoots could be induced to form by appropriate growth regulator treatments.

Hackett and Anderson (1967) got either single shoots from carnation shoot apices, or else a proliferative tissue from which shoots were later regenerated. Walkey and Woolfitt (1968) reported a similar kind of direct or indirect shoot proliferation from Nicotiana rustica shoot tips. Vine and Jones (1969) were able to transfer large shoot tips of hop (Humulus) to culture, but shoots only rooted, and showed a high propensity for callus formation. Reports of plant multiplication using conventional shoot culture methods began to appear in the next decade.

Haramaki (1971) described the rapid multiplication of Gloxinia by shoot culture and by 1972 several reports of successful micropropagation by this method had appeared (Adams, 1972; Haramaki and Murashige, 1972). Since then the number of papers on shoot culture published annually has increased dramatically and the method has been utilised increasingly for commercial plant propagation. Factors which have influenced the choice of shoot culture for practical micropropagation have been:

• the way in which the method can be applied to a wide range of different plant species, using the same principles and basic methods;

• the possibility of obtaining simultaneous virus control;

• a general uniformity and 'trueness to type' of the regenerated plants;

• the relatively high rates of propagation which is possible in many species.

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