There are multiple methodologies being followed for different types of plant cell/tissue culture techniques. The current know how in this area is such; it is not only possible to culture free cells, genetically alter them, induce cell divisions, and elicit secondary metabolite production; plant cells cultured from plants with desired combination of traits may be somatically fused to form artificial hybrids, plantlets regenerated, and whole plants could be raised from it. Cell culture allows study of plant cell metabolism and effect of various substances on cellular responses and also enables clonal propagation of genetically identical plants. Thus, depending on the purpose, cultures may be categorized as organ, tissue, and cell cultures.

Organ Culture

Aseptic culturing of isolated plant organs or those obtained from callus differentiation is called organ culture. It is possible to culture virtually any plant part such as root, leaf, stem, flower, seed, and cotyledon. These are cultured for plantlet regeneration or to induce callus formation. Cultured organs can be made to generate secondary metabolites in higher yield than intact plants, for example, leaf cultures of D. purpurea and D. lanata. In cases as in leaf cultures of Vinca and Rauwolfia, a variety of constituents not found in intact plants are produced. Similarly, leaf and root cultures of C. ledgeriana accumulated quinine and quinidine not found in cell suspension cultures. Root cells genetically transformed through bacteria such as Agrobacterium tumefaciens, when grown in a medium devoid of growth hormones, give rise to numerous fine hairy roots. Such transformed hairy root cultures of several medicinal plants reportedly accumulate much higher yields of normal secondary metabolites as well as compounds not associated with intact plants. For example, Solasodine content in S. laciniatum cultures was four times over intact plants. Hairy root cultures of P. ginseng reported more effective production of ginsenosides over ordinary root cultures. Similarly valepotriate yield was four times over whole roots in V. officinalis hairy root cultures. Ginseng hairy root cultures have reportedly transformed added digitoxigenin to novel stearate, palmitate, and myristate esters.

Tissue Culture

Excised plant-tissues such as root tips, shoot tips, leaf primordia, ovaries from pollinated or unpollinated flowers, ovules, anther, embryos, nucleus and endosperm are cultured in vitro for plant cell physiological/morphological studies, clonal propagation, plant hybridization, and in plant breeding programs.

Cultured plant-tissues are grown on hormone-controlled media to induce callus formation. Callus actually refers to wounded tissue that an intact plant produces when there is injury. In tissue culture, proliferating mass of unorganized/undifferentiated parenchymatous cells arising from an explant is called callus. Such callus cultures could be maintained for prolonged periods by repeated sub-culturing. They may thus be made to form an extended area of tissue or form a friable mass, which can be dispersed into a suspension culture.

Callus cultures may be utilized for plant regeneration, preparation of single-cell suspensions, protoplast isolation, and in genetic transformation studies. Callus derived cultures from medicinal plants are being much investigated and form an important source of plant secondary metabolites. Explant or the plant part that generates the callus is crucial in determining the nature of secondary metabolites formed in culture.

Single-Cell Culture

Culturing single cells derived from callus cultures or by mechanical or enzymatic isolation from plant material is called cell culture. Since they are totipotent and carry the required genetic information, by hormonal manipulation they may be induced to form roots, shoots, and thus regenerate into a complete plant in culture. Alternatively, when isolated single cells are placed in a conditioned medium, they undergo repeated division, multiply, and result in callus formation. When single cells are cultured in a conditioned liquid medium, they are called suspension cultures. In this way it is possible to culture them in large volume bioreactors similar to micro organisms and they can be maintained in continuous culture for the production of a range of high value, low volume phytochemicals. Unlike microbes, cell proliferation rates are much lower and they have a tendency to grow as aggregates. Also being fragile, dispersing aggregates in large volume liquid cultures requires special low-shear mixing techniques to prevent cell rupture. Free cell suspension cultures can be grown using the following techniques:

Batch Culture

It is a suspension culture of smaller volumes ranging from 100–250 ml, being agitated continuously to break aggregates of cells. Cells grow and multiply in medium constituted of limited amount of nutrients, after a lag period of cell acclimatization. Following an exponential growth and division phase, due to exhaustion of a nutrient, they fall into the stationary growth phase. As no further medium is added in each batch of cultured cells, it is a “closed” system and is generally used for initiating single-cell cultures. Such cultures may be subcultured by transferring aliquots into fresh medium. Such repeated culturing may generate a fine suspension of cells.

Continuous Culture

Here cells are grown in suspension in media that is limited in quantity with respect to one nutrient. Since cell growth is limited by this nutrient, before its exhaustion, fresh medium is added to this limiting nutrient. Growth of cells in culture is thus continuous. When the old medium is withdrawn, if cells are separated and added back to culture, it is a “closed continuous culture.” However, with addition of medium, there is withdrawal of suspension, without adding back cells, it is an “open continuous culture.” Here the medium input and culture harvest are so balanced that cells are in a constant submaximal growth rate continuously. When media addition is determined by constant level of a particular nutrient, the culture is a “chemostat.” If, however, media addition is intermittent, as and when the turbidity of the culture is enhanced due to an increase in number of steadily growing cells, it is called a “turbidostat” culture.

Protoplast Culture

Protoplasts are cells without cell walls. Though they may be isolated directly from any plant part such as young leaf, root, stem, seedling, pollen grains, petals and embryo sac, leaf mesophyll cells and cells from suspension culture are convenient for protoplast isolation. Several enzymatic and mechanical means are employed for stripping the cells of their cell wall. Such protoplasts, almost spherical in shape may be cultured by plating them on agar. The pinocytic activity, that is, the ability to take up extracellular material and totipotency make them ideal research tools for both fundamental studies in cell biology and applications based on somatic hybridization and genetic engineering.