3. Initiation and establishment of cell suspension cultures

3.1. Callus cultures

When an organ of a plant is damaged a wound repair response is induced to bring about the repair of the damaged portion. This response is associated with the induction of division in the undamaged cells adjacent to the lesion, thus sealing of the wound. If, however, wounding is followed by the aseptic culture of the damaged region on a defined medium, the initial cell division response can be stimulated and induced to continue indefinitely through the exogenous influence of the chemical constitution of the culture medium. The result is a continually-dividing mass of cells without any significant differentiation and organization and this proliferated mass of cell aggregate is called callus. The first step to establish cell suspension cultures is to raise callus from any explants of the plant. To maximize the production of a particular compound, it is desirable to initiate the callus from the plant part that is known to be high producer.

Calli are generally grown on medium solidified with gelling agents like, agar, gelrite, agarose, in Petri-dishes, glass test-tubes or extra-wide necked Erlenmeyer flasks. In morphological terms it can vary extensively, ranging from being very hard/compact and green or light green in color, where the cells have extensive and strong cell to cell contact, to being ‘friable' where the callus consists of small, disintegrating aggregates of poorly-associated cells and has brownish or creamy appearance (Figure11.1). Friable callus is most demanded since it shows fast and uniform growth of cells and is highly suitable to initiate cell suspension cultures. Callus morphology is explants and species dependent but can be altered by the modification of plant growth regulators in the medium.

The callus cultures shows inherent degree of heterogeneity and this may be due to their size and nature, unidirectional supply of nutrient medium (below the callus) and gases and light (predominantly from above). The heterogeneity may be disadvantageous in uniform production of cell biomass but may be useful in the developmental responses of the callus like, shoot regeneration.

Figure 11.1: Callus cultures- A. Hard and compact callus; B. Friable and brown callus