05 Growing Process

Growing Process¶

Step 1: Wash your hands with soap and then spray ethanol.
Step 2: Sterilize the work area with ethanol alcohol. Carefully light a Bunsen burner and place the substrate and culture in a sterile zone near the flame.
Step 3: Open bottle and put the lid on the table.
Step 4: Open the tube, throw the pure culture into the bottle and close the bottle immediately.

Do not touch the pure culture

Step 5: Move the culture to the middle of the substrate by slightly shaking of the bottle.
Step 6: Keep the bottle in your living room or bedroom. Far from the window (avoid direct sunshine). Do not keep it in totally dark (light is necessary for fruiting).

The best temperature for fruiting is between 20℃ - 30 ℃

After five days, the mycelium began to spread through the substrate, marking one of my first mushroom cultivation experiments. This week, I'll be cultivating a new batch of fungi using the agar plate. Then, I will compare the growth time and fruiting of pure cultures grown on the substrate with those grown on an agar medium. The liquid format offers flexibility for sample distribution.

I've chosen to focus on fungi native to tropical and humid environments for this initial research because the bacteria and the medium to expand the lifespan take much time to deliver to Spain.

Medium Preparation¶

I selected three main nutrients: malt extract, potato starch, and yeast to create the medium for the fungus pure culture. All of these were placed in a petri dish with agar.

After sterilizing the materials, it's crucial to ensure that the work surface is also sterilized. To do this, use ethyl alcohol and place a Bunsen burner in the center to create a sterile working environment. Then, place all the materials you'll be using around the burner, as close to the flame as possible.

Since I used three nutrients, I first created the agar base and then added the nutrients. For the malt extract and potato starch, I followed two different recipes (see the datasheet). During the process, I made a mistake that turned out to be interesting: I didn't add agar to the malt extract, and it's the culture that's growing the fastest. However, I want to conduct another test to validate whether this is due to the nutrient itself or if the fungus simply grows faster in liquid cultures. On the other hand, I conducted a test with an agar and cotton base, which was also interesting because the mycelium is growing quite quickly on the fabric.

Process step by step¶

Step 1: Sterilize all tools with alcohol and dry thoroughly. Then weigh the nutrients and mix them with water in the bottle until the nutrient is dissolved.

Step 2: If you are working with different recipes, you should label the bottles to differentiate the nutrients. In my case, I labeled the medium containing agar with tape.

Step 3: Place water in the pressure cooker with water (¾). Place the bottles and all the materials you are going to use in a bag and leave the lid of the bottles a little open.

Important: When placing the medium in the bottle, leave the lid slightly open to prevent it from exploding.

Step 4: Close the bag by rolling it up, then put it into a pressure cooker for 30 minutes.

Step 5: After 30 minutes, remove the instruments and materials from the pressure cooker and place them in the sterilized area as close as possible to the burner. Wait 10 minutes while the medium cools down so you can use it.

Step 6: Place the sterilized petri dishes and medium in the sterilized area. Then bring the bottle close to the burner to open it and prevent contamination, place the mouth of the bottle in the fire, keep the bottle close to the fire, open a little of the lid of the petri dish, and carefully place the medium.

Step 7: Then open the lid of the petri dish a little in the direction of the fire so that the medium solidifies and does not generate steam stains on the lid.

Step 8: When the medium has solidified, add the pure culture.

Step 9: Identify the sample on the edge with the date, nutrient name, and culture, and seal the lid with paraffin.

Step 10: Finally, place the samples in the incubator in the temperature range of 20-30°C. Monitor the samples daily.

If you see any dark spots, the culture may be contaminated. In that case, remove them from the incubator, document the sample, and discard it.

Contaminated samples¶

Experiments datasheet¶

Bioluminicense¶

In this image, you can see on the right the faint bioluminescence that began to emit the growing kit on the substrate. This is a cell phone photo and was not edited. The light emission is more evident when we look at the sample live.