How to prepare Agar plates (Advanced)

The Importance and Challenges of Using Agar in Mushroom Cultivation

Mushroom cultivation is a fascinating and rewarding endeavor, but it often requires a deep understanding of various techniques and materials, with agar playing a pivotal role in the process. Agar, a gel-like substance derived from seaweed, is indispensable in the world of mycology for several reasons, yet it also presents certain challenges that cultivators must overcome.

Importance of Agar in Mushroom Cultivation:

1. Isolation of Mushroom Strains: Agar is primarily used for isolating and maintaining pure mushroom strains. This is crucial for preserving the genetic characteristics of a particular mushroom variety and ensuring consistent yields and quality.
2. Spore Germination: When working with spores, agar provides a solid medium for spore germination and the formation of mycelium. This step is fundamental for propagating mushroom cultures.
3. Cloning: Agar allows for the cloning of desirable mushroom strains by transferring a small piece of mycelium onto fresh agar plates. This technique is essential for scaling up production and maintaining desired traits.
4. Contaminant Control: Agar’s solid nature helps prevent contaminants from infiltrating mushroom cultures. It acts as a barrier, protecting the developing mycelium from unwanted bacteria or molds.
5. Research and Experimentation: Agar plates are ideal for conducting experiments and research in mycology, making it possible to study various aspects of mushroom growth and development.

Challenges of Using Agar in Mushroom Cultivation:

1. Contamination: While agar helps prevent contamination, it is not foolproof. Contaminants can still find their way into cultures, leading to failed experiments or contaminated mushroom substrates.
2. Sterility: Maintaining a sterile environment when working with agar is challenging and requires rigorous protocols, including sterilizing equipment and working in a laminar flow hood or glove box.
3. Cost: Agar can be relatively expensive, especially for larger-scale operations. The cost of agar, along with other laboratory supplies, can add up quickly.
4. Skills and Knowledge: Working with agar requires a good understanding of sterile technique and microbiology. Novice cultivators may face a steep learning curve.
5. Environmental Impact: The production of agar can have environmental implications, as it is sourced from seaweed. Sustainable agar sourcing practices are essential to mitigate this issue. Petri dishes are mostly made of plastics, you can cut down on plastic waste by using either reusable pp5 or glass dishes.

In conclusion, agar is an invaluable tool in mushroom cultivation, facilitating the isolation, propagation, and research of mushroom strains. Its use is crucial for maintaining genetic purity and advancing the field of mycology. However, it also presents challenges related to contamination, cost, and the need for a high level of skill and knowledge. Cultivators must strike a balance between its importance and the potential difficulties it brings to ensure successful mushroom cultivation.

 

Potato Dextrose Agar (PDA) is a commonly used agar medium for growing fungi and culturing mushroom mycelium. Here’s a recipe for preparing PDA agar:

Ingredients:

• 200 grams of peeled and diced potatoes
• 20 grams of dextrose (glucose)
• 15-20 grams of agar-agar powder (depends on manufacturer)
• 1 liter of distilled or deionized water

Equipment:

• Saucepan
• Blender or food processor
• Beaker or glass measuring cup
• Stirring rod
• pH meter and pH adjusting solutions (if needed)
• Autoclave or pressure cooker
• Sterilized Petri dishes or containers
• Sterile still air box or laminar flow hood
• Gloves and a face mask

Procedure:

1. Potato Preparation:

1.1. Peel and dice the potatoes into small pieces.

1.2. Rinse the diced potatoes thoroughly under cold running water to remove excess starch.

2. Potato Extraction:

2.1. Place the diced potatoes in a saucepan and add enough distilled or deionized water to cover them.

2.2. Bring the water to a boil and simmer the potatoes for about 30 minutes or until they become soft.

2.3. Filter the solution with a coffee or tea filter so you get a mostly clear liquid.

3. Agar Solution Preparation:

3.1. Add 15-20 grams of agar-agar powder to solution and mix it well to combine.

3.2. Adjust the volume of the mixture to 1 liter with distilled or deionized water if needed.

3.3. Add 20 grams of dextrose (glucose) to the mixture and stir until it is completely dissolved and heat until boiling. It’s important to boil the solution to                make sure it will solidify later on.

4. Sterilization:

4.1. Cover the beaker or glass bottle containing the agar mixture with aluminum foil to prevent contamination during sterilization, don’t seal your container          to prevent it from exploding while autoclaving.

4.2. Place the covered beaker or bottle in an autoclave or pressure cooker.

4.3. Sterilize the PDA agar at 121°C (250°F) and 15 PSI for 15-25 minutes in an autoclave, or follow the sterilization instructions for your specific pressure             cooker.

4.4 Let the pressure cooker/autoclave cool down slowly without venting, best to cool it down in front of a flow hood.

4.4. Allow the agar to cool to around 50°C (122°F) before proceeding.

5. Pouring Agar:

5.1. Set up your still air box or laminar flow hood in a clean, sterile environment.

5.2. Wear gloves and a face mask inside the controlled environment to minimize contamination risk.

5.3. Place the sterilized Petri dishes or containers inside the controlled environment.

5.4. Carefully remove the aluminum foil from the beaker or cup containing the sterilized agar.

5.5. In the controlled environment, pour the agar into the Petri dishes, covering the bottom to a depth of approximately 3-4 mm. Tilt the dishes slightly to evenly distribute the agar.

5.6. Allow the agar to solidify by leaving the Petri dishes undisturbed for about an hour.

6. Storage:

6.1. Once the agar has solidified, seal the dishes with parafilm or grafting tape, store the Petri dishes or containers in a cool, dry place until you’re ready to use them for inoculation or other experiments.

This PDA agar recipe provides a suitable medium for culturing various fungi and mushroom mycelium. Proper sterilization and aseptic technique are crucial to ensure the agar remains free from contaminants during preparation and use.

 

Here are some other recipes you can try:

related article:

How to work with Agar plates