Callus is group of parenchymatic nature of cell which have no defined function. callus culture is technique in which mass of undifferentiated plant cell are grown in artificial media in laboratory.
Imagine we want to study how plants grow in space or create plants with super resistance to diseases. But growing full plants can be slow, space-consuming, and hard to control. This is where callus cultures come in.
They allow us to grow plant cells in a lab under controlled conditions, making it easier to understand plant growth and manipulate plants for research or agricultural purposes.
These techniques are at the core of plant biotechnology, helping us produce medicines, improve crops, and conserve endangered species.
What is Callus Culture?
A callus is a mass of undifferentiated plant cells that grow on a solid medium, often initiated by wounding a plant or using specific plant hormones.
In callus culture, we take small pieces of plant tissue and grow them on a nutrient-rich medium in a lab. These cells multiply, forming a clump, which we can manipulate for further studies or regeneration into a full plant.
Explants tissue can be : roots, stem, leaves, flowers etc.
Factors important for the formation –
- Hormons – Auxin alone, Cytokinin alone or Both
- Genotype,
- Composition of nutrient medium,
- Physical growth factors (light etc.)
- Types of Sugar source (glucose or sucrose)
Different characteristics of callus
Callus tissue induced form different plant species may be different in structure and growth habit :
- White or colored
- Soft (watery) or hard
- Friable (easy to separate)
- Compact
Initiation and maintenance of Callus Culture
Starting a callus culture involves the following steps:
- Wash the seeds using water with few drop of detergent in a small vessel/container/beaker and shake.
- Submerge the seed into 70% alcohol for around 30-60 second and then discard the alcohol.
- Transfer the seeds to a flask or beaker containing 20-40% sodium hypochlorite for 15-20 min. Then rinse the seeds for 4-5 times with sterile distilled water.
- Transfer 2-3 seeds/flask on the surface of MS agar medium without growth regulators.
- Incubate the culture at 250C under photoperiod of 16 light (~1000 lux intensity) for 1-2 week.
- Collect the seedling aspectically (about 2 week old) and place them into sterile petri dish and prepare the explants.
- Excise shoot apex and inoculate them into nutrient medium (containing 1-2 mg/l Auxin)
- Incubate the cultures in dark at 250C for about 3-4 week.
- Check the growth and if require cut small pieces of callus and subculture on the same fresh medium.
Growth of Callus
Culturing of regularily new medium exhibit an S-shaped or sigmoid pattern of growth during each passae.
- Lag phase : cell prepare to divide
- Exponential phase : Highest cell division
- Linear phase: Slow division but increase cell expansion
- Deceleration phase : decrease division and elongation.
- Stationary phase : cell remain constant
Growth can be monitored by – Weigh measurements, color change or any irregularity, Mitotic index to reduce sample error.
Maintenance of Callus Culture
- Subculturing : Periodically transfer portions of callus tissue onto fresh nutrient media to promote continuous growth. Prevent overgrowth or browning by subculturing at optimal intervals.
- Monitoring: Regularly inspect cultures for contamination, changes in color, texture, and growth patterns.
- Adjustment of Growth Regulators : Modify concentrations of auxins and cytokinins in the medium to control growth and induce specific responses (e.g., organogenesis or embryogenesis)
Advantages of Callus Cultures
- Mass Production: Large quantities of undifferentiated cells can be generated quickly.
- Genetic Engineering: Callus tissue can be manipulated easily for introducing new genes, allowing the development of genetically modified plants with desirable traits (disease resistance, increased yield, etc.).
- Regeneration of Plants: Under the right hormonal conditions, a callus can regenerate into an entire plant, a process known as organogenesis.
- Conservation: Callus cultures help preserve rare and endangered plant species by allowing their propagation in vitro.
Limitations of Callus Cultures
- Loss of Differentiation: Over time, callus cells may lose their ability to differentiate into specific tissues (e.g., roots, shoots).
- High Contamination Risk: Cultures are prone to contamination by bacteria or fungi, especially during the transfer and sub-culturing steps.
- Cost: The setup and maintenance of sterile culture environments can be expensive.
What is Suspension Culture?
Suspension culture involves growing plant cells in a liquid medium. Unlike callus culture, where cells are anchored on a solid surface, suspension cultures have freely floating cells or small clusters in a constantly shaken or stirred liquid environment, promoting cell division and uniform growth.
Types of Suspension Culture
- Batch Suspension Culture: Cells grow in a liquid medium until nutrients finish, then the culture is harvested.
- Continuous Suspension Culture: Fresh nutrients are constantly added, and cells are continuously grown and harvested, making it more efficient for long-term studies.
Suspension Culture Advantages:
- Uniform Exposure: Cells in a suspension culture have uniform access to nutrients and oxygen, leading to consistent and rapid growth.
- Scalability: It is easier to scale suspension cultures for large-scale production of plant metabolites, bioactive compounds, or cell mass for research.
- Automation: Suspension cultures can be automated for continuous operation, making them ideal for industrial purposes.
Suspension Culture Limitations:
- Fragility of Cells: Cells in suspension are more sensitive to mechanical damage from agitation, which can affect their growth and viability.
- Difficulty in Regenerating Plants: It is harder to regenerate whole plants from suspension cultures compared to callus cultures.
- Maintaining Homogeneity: Cells in suspension can grow in clusters, making it difficult to maintain a uniform population for experiments.