Cultures from embryogenic callus (Stage I). Suspension cultures can sometimes be initiated from embryogenic callus tissue, and the cells still retain the capacity to regenerate somatic embryos freely. Obtaining such cultures is not always a simple matter, for the auxin levels that are often used to promote cell dispersion may result in the loss of morphogenic capability. Embryogenic cell suspensions are most commonly initiated from embryogenic callus that is placed in liquid medium on a shaker. Vasil and Vasil (1981a,b) and Lu and Vasil (1981a,b) have reported producing cultures of this type from pearl millet and guinea grass respectively. Suspensions were initiated and subcultured in MS medium containing 1-2.5 mg/l 2.4-D and 2.5-5% coconut milk, and came to be composed of a mixture of embryogenic cells (small, highly cytoplasmic and often containing starch) and non-embryogenic cells (large and vacuolated). Embryoids were induced to develop into somatic seedlings when plated onto an agar medium without growth regulators, or with lower levels of auxin than used at the previous stage.
Cultures from non-embryo genic sources. Embryogenesis can be induced in cell suspensions of some plants when the cultures are produced from non-morphogenic callus and have been maintained without morphogenesis for one or more transfers. Induction occurs most readily in recently isolated suspensions and usually becomes much less probable with increasing culture age. Loss of regenerative ability is often associated with the appearance of some cells with abnormal chromosome numbers, but it can also be due to culture on an inappropriate medium.
Embryogenesis in suspension cultures seems to require media at Stage I and Stage II with similar compositions to those necessary for somatic embryo formation in callus cultures. Somatic embryos can be formed in suspension cultures in very large numbers. Reinert et al. (1971) demonstrated that the continued capacity of carrot cell suspensions to form embryos depended on an adequate supply of nitrogen. Embryogenesis ceased on a medium containing little nitrogen, but it was re-induced for several transfers after the culture was returned to a high-nitrogen medium.
In a few kinds of plants it is possible to induce embryogenesis in previously unorganised suspension cultures. Success is so far recorded only in members of the families Apiaceae (Umbelliferae), Cruciferae and Scrophulariaceae. This is not therefore a method of propagation which can be readily utilised. There is a greater chance of obtaining an embryogenic suspension culture from embryogenic callus.
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