Preparation
Preparing the environment before inoculating spores is a critical step in the success of any cultivation project. A sterile and controlled environment is essential to prevent contamination from unwanted microorganisms that can compete with or outgrow the mushroom spores.
By creating an ideal setting, you promote healthy spore germination and mycelium development, while minimizing external factors that could impede growth, such as fluctuating temperatures or humidity levels. Preparing the environment also helps reduce the risk of contamination, allowing for faster and more reliable results.
Furthermore, a well-prepared workspace ensures consistency, leading to reproducible outcomes in mushroom cultivation or other fungal projects. Maintaining sterilized tools and surfaces and controlling environmental conditions is key to achieving optimal growth and ultimately achieving the desired results.
TOP TIP: Before preparing your space, make sure your substrate, spores, agar, etc. are not in the room. Stray disinfectant can upset your project by killing spores or breaking down air filters.
How to Prep your Space
There are many different types of disinfectant, and if you're in a pinch soap and water will do. However, the best disinfectant to use would be 70% Isopropyl Alcohol Spray.
Simply spray or wipe down surfaces like countertops, tools, and containers, and it’s also useful for sterilizing your hands before handling spores or substrates.
Alternatively, bleach diluted with water (10% bleach to 90% water) can be used on non-porous surfaces like countertops, floors, and tools. Be sure to work in a well-ventilated area, as bleach produces strong fumes. Avoid using bleach on porous materials, as it can be less effective and may damage the surface.
Always allow the area to air out prior to introducing your project or tools into the room.
Getting Started with Agar
Agar work can appear intimidating, but it doesn't have to be. This is not intended as a complete guide, but rather the foundation you can use to build your knowledge and skill up!
Agar is best used in lab-like conditions. Your spare bedroom or airing cupboard may work for your grow kits, but agar will pick up any little microbe in the air and grow that instead of your spores. Therefore, we would strongly recommend the use of a Flow Hood, or Still Air Box (SAB). Please note that a SAB is not the equivalent to a Flow Hood and will not offer the same amount of protection.
Still Air Box
Cost-effective- cheaper and can be made with basic materials.
Simple to Use- straightforward set-up, no prior experience or knowledge needed.
Portable- smaller and doesn't require external power.
Limited Airflow Control- cannot filter out airborne contaminants as effectively.
Limited by User Ability- success depends on the users ability to work within it without introducing contaminants.
Flow Hood
Superior Contamination Control- constant flow of air ensures airborne contaminants are removed.
Efficiency- high-volume work is easily performed, significantly reducing contamination risk.
Cost- more expensive.
Space- takes up more space, needing a dedicated area or room
Maintenance- require filter changes and user ensuring the system is functioning properly
To begin your studies with Agar, you will need a few tools prior to getting started. As mentioned above, a SAB or Flow Hood is useful to have.
You will need:
Agar Dish
Spores or Mycelium
Sterile Tools: inoculation hoop, scalpel, needle, etc.
70% Isopropyl Alcohol or Bleach Solution
Parafilm of Tape
You can purchase spores in swab, print or syringe form for us which can be used with this method. Swabs and prints are best as the added water in the solution can disturb the Agar. You can also take samples from mature fruits.
Inoculating your Agar
If you have a Flow Hood or SAB, be sure to set it up prior to these steps to ensure the environment and equipment remains sterile.
- Sterilize your inoculation tools (scalpel, needle, or loop) via autoclave. You can heat them with a flame, however this method will not ensure a contamination-free project. Using new, disposable, hoops, scalpels or loops would be safer.
- For liquid spores, drop a small amount of spore solution from a syringe onto the agar. If you're using a swab or print, lightly press spores in swab form into the agar and move it across the surface in a zig-zag formation. For live tissue, cut a small piece of mushroom tissue and place it in the middle of the agar plate.
- Quickly cover the Petri dish after the inoculation and seal the edges of the dish with Parafilm or tape to keep the contaminants out. Label the dish with the species, strain and date of inoculation.
- Incubate the petri dishes in a dim or dark area with a stable temperature of 20-25c. Ensure you avoid direct sunlight or extreme temperatures.
- Check regularly for signs of contamination (e.g., green, black, or slimy spots). Healthy mycelium appears white and grows outwards from the inoculation point. If contamination does occur, ditch the dish or cut out clean sections and transfer to a new agar plate.
- One the mycelium has grown to almost reach the edges of the plate, transfer it to new plates to isolate clean cultures or proceed to grain spore inoculation.
Healthy white mycelium in an agar plate.
Swiping the spores throughout the agar like this will encourage growth.
