Plant Callus Culture: Definition, Types, and Procedure

Define Callus and Describe Procedure For Callus Culture

A callus culture is a mass of irregular undifferentiated parenchymatous and totipotent cells. These cells have the ability to divide and differentiate to form an entire organism. It is formed in culture media due to active mitotic division of the cells of the explant. Callus culturing is used in the production of secondary metabolites for plant regeneration.

Types of Callus Culture

• Compact Callus
• Friable Callus

1) Compact Callus

It shows densely aggregated cells.

2) Friable Callus

It shows loosely associated cells, and the callus becomes soft and breaks easily.

Stages of Callus Culture

There are three stages of callus culture:

1) Induction

Cells in the explant dedifferentiate and begin to divide.

2) Proliferative Stage

Rapid cell division occurs.

3) Morphogenesis Stage

Differentiation and formation of organized structures, specifically processes that lead to plant regeneration from somatic cells.

Procedure for Callus Culture

1. Selection and collection of explant material
2. Preparation of culture media
3. Surface sterilization of explant material
4. Preparation of explant
5. Inoculation of explant in culture medium
6. Incubation of culture
7. Subculturing
8. Regeneration of plant from callus

1) Selection and Collection of Explant Material

Any kind of permanent but living tissues, such as parenchyma, collenchyma, cortical tissue, pith cells, phloem cells, cambium cells, or meristematic tissue, may take part in callus formation.

2) Preparation Of Culture Media

The most common nutrient media used for callus culture is MS media, which is sterilized by an autoclave before use.

3) Surface Sterilization

In this step, microbial contaminations present on the surface of the explant material are killed by using different chemical agents:

• Wash explant material with tap water. If any waxy substances are present, treat with liquid detergent for 10–15 minutes.
• Treat with 70% Ethanol for 1–2 minutes.
• Now treat it with a chemical sterilizing agent (sodium hypochlorite).
• Wash with distilled water.

4) Preparation of Explant

Surface-sterilized explant material is cut into small pieces with a sterilized knife.

5) Inoculation of Explant in Culture Medium

The explants are finally transferred aseptically onto a suitable nutrient medium solidified with agar.

6) Incubation

Incubation of the culture under controlled physical conditions, such as temperature, light, and humidity, is necessary for the proper initiation of callus tissue. Here, explant cells utilize nutrients from the media, grow, multiply, and are converted into a mass of cells called callus.

7) Subculturing

This involves the maintenance of callus culture. Subculture is the process by which the tissue or explant is first subdivided and then transferred into fresh culture medium.

8) Regeneration Of Plant from Callus

Plantlets can be obtained from callus culture cells by two different ways:

• Organogenesis
• Embryogenesis