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How to Isolate Fungi From Mixed Flora

Fungi must be isolated from the contaminating influence of the wild mixed flora of the fungal habitat. Once separated from other lifeforms that overgrow, weaken, or kill the fungi, it's ready to be grown as pure strains in a controlled laboratory evironment. Some of the competing flora grow faster than the fungi and devour its food supply. The mixed flora, called indigenous microbiota by scientists, include protists, bacteria, other non-desirable species of fungi and yeast.

Things You'll Need

Clean-room
Inoculation box
Air purifier
Latex gloves
Surgical mask
Isopropyl alcohol
Agar mix
Water
Autoclavable container
Petri dishes
Scalpel
Pressure cooker
Wild fungi population
Plastic bag
Iodine solution
Alcohol lamp

Instructions

- 1
  Set up and outfit a clean-room to grow the fungi you want to isolate and purify from the mixed flora contaminents of its outdoor environment. Choose a freshly painted room with little air circulation. Wear clean clothes and shower before working with fungi cultures. Install an air purifier. Prevent contamination by wearing latex gloves and a mask. Spray your gloves frequently with isopropyl alcohol. Keep the level of mixed flora organisms low in the room.
- 2
  Prepare the agar media to grow the fungi during the isolation and purification process. Follow the directions on the container to dissolve dry agar in water in an autoclavable container. Stack petri dishes, scalpels and the agar inside a pressure cooker. Sterilze by cooking for 25 minutes at 15 psi. Cool the cooker and place the dishes and tools in an inoculation box with a glass lid and holes in the front for your hands. Pour the agar into the dishes while it's hot. Fill them half way.
- 3
  Locate a fungi population you wish to isolate. Select a clean fruiting body, known as a carpophore. Put on sterile gloves, collect the carpophore and put it in a clean plastic bag. Take it to the lab and put it on a sterile surface. Disinfect the carpophore skin with an iodine and alcohol solution. Cut out tiny sections from the interior after peeling back the skin. Place the pieces in the center of the petri dishes.
- 4
  Flame the scalpel with a lab alcohol lamp after several cuttings to sterilize it. Adjust the temperature and humidity level to the specific type of fungi being isolated. Observe the fungi as it grows out into the medium from the chunk. Cut out and discard any areas colonized by competing fungi. Dump the culture if yeast, puddles of bacteria or black and green mold take over. Identify and purge the fungi cultures of any unwanted mixed flora.
- 5
  Continue to isolate the fungi you want to grow from the contaminant flora. Cut out triangles of agar overgrown with pure fungi culture in the petri dishes. Pick sections far from any growing contaminants. Stab the triangles with the scalpel and transfer each piece to a new dish of agar. Do this until only pure isolated fungi cultures are observed growing.

Nature