Autoclave in Plant Tissue Culture

The autoclave is a critical, indispensable piece of equipment in a plant tissue culture laboratory.Its primary function is to ensure the absolutely necessary sterile (aseptic) conditions required for successful in vitro plant growth.

It achieves sterilization using the principle of moist heat sterilization—high-pressure steam.

The Autoclave’s Role in Plant Tissue Culture

The autoclave is used to sterilize virtually everything that will come into contact with the delicate plant tissue and its growth environment, including:

1. Culture Media: This is the most important application. The nutrient-rich medium (containing salts, sugar, vitamins, etc.) is a perfect environment for microorganisms (bacteria, fungi, and spores).Autoclaving destroys these contaminants, typically at 121^◦ C(250^◦ F) and a pressure of 15-20 psi (1.05-1.4kg/cm² ) for a volume-dependent time (~15-20 minutes for small volumes).
2. Culture Vessels: Glassware (flasks, test tubes, jars, Petri dishes) used to contain the culture medium and plant tissue.
3. Tools and Equipment: Metallic instruments like forceps, scalpels, and spatulas.
4. Water and Solutions: Sterile water or certain heat-stable stock solutions.

Mechanism of Sterilization

The key to the autoclave’s effectiveness lies in the use of steam under pressure:

• Pressure increases boiling point: By increasing the pressure inside the chamber, the boiling point of water is raised from 100^◦ C to 121^◦ C.
• Moist Heat Kills: The high-temperature steam rapidly transfers heat to the items being sterilized.The moisture in the steam causes the coagulation (denaturing) of essential proteins within the microbial cells, leading to their irreversible destruction, including heat-resistant spores.

Important Consideration: Thermolabile Components

While the autoclave is the gold standard for sterilization, certain components of the culture medium—like some vitamins, hormones (plant growth regulators), and certain carbohydrates—are thermolabile (heat-sensitive).

• Handling Thermolabile Components: These components would decompose or lose their effectiveness if autoclaved.
• Alternative Sterilization: They are typically prepared as separate stock solutions and sterilized using membrane filtration (passing the solution through a sterile filter with a very small pore size, typically and then added aseptically to the cooled, already-autoclaved main medium.