Tissue culture Techniques

The key word behind the success of tissue culture is the “totipotency”

The various techniques or principles of plant tissue culture are as listed below

  • Callus culture
  • Meristem culture
  • Organ culture
  • Protoplast culture

Callus Culture

The callus is a mass of highly vacuolated, unorganized cells resulting as a consequence of wounding in plants, and in tissue culture with the use of sophisticated techniques.

A cell, may be from pollen, anther, bud, fruit, leaf or shoot apex etc. when cultured on a suitable growth medium under aseptic, controlled conditions, differentiate and multiply to form callus. This callus by induced re-differentiation can give rise to embryoids. Embryoids develop into plantlets and later on in whole viable plant.

Principles of Callus Culture

  • Aseptic preparation of plant material
  • Selection of suitable growth medium (auxin : cytokinin ratio is important to consider)
  • Selection of suitable controlled physical growth conditions for incubation

Meristem Culture

A meristem is a group of undifferentiated plant cells (found at growth tips) which can undergo divisions to form all types of tissues. Generally explant used is shiny dome shaped structure of length less than 0.1mm with one or two pairs of youngest leaf primordia.

Meristematic tissue are isolated and cultivated on a suitable growth medium under aseptic conditions. Meristem forms callus at its cut end on which a large number of shoot primordia develop. These primordia develop into multiple shoots, which after rooting produce small plants bearing 5 or 6 leaves. (The shoots grow out directly form excised shoot tip cultured. In shoot tip culture the explant used is meristem along with primordial and developing leaves and adjacent stem tissues.)

Applications of meristem culture

  • Virus and parasite elimination – The resulting plantlets are often free of viruses and parasites. Meristems are devoid of viruses. There are certain reasons behind this.
    • Vascular system is absent in meristem. Vascular system has prime importance in the travelling of viruses through the body.
    • Meristematic cells are actively dividing and have high metabolic activity. These factors have negative impact on virus multiplication.
    • Shoot apex have high endogenous auxin level, which act as inhibitory factor for virus multiplication.
  • Used in propagation of haploid plants.
  • Storing genetic resources of seed producing plants of heterozygous nature.
  • Used in micropropagation

Organ culture

In organ culture specific organ is excised and cultured on a suitable growth nutrient medium under aseptic and controlled physical conditions. Here, the particular organ retains its characteristic structure and features and continue to grow as usual unlike in callus culture where organization of intact tissue is lost.

It includes anther culture, pollen culture, culture of embryo, ovary, root, shoot, floral buds, fruit, seed, leaf primordia etc.