Barbara Pollini, Simeon Goodwin, Eli Brouwer, Antoni Gandia (AD), Marieke Broertjes

Agar Petri Dish Preparation for Growing Mushrooms

Agar medium recipes

The first phase of mushroom cultivation is the isolation of a pure culture of mycelium. These cultures are grown in petri dishes, with a medium (food) for the mycelium to digest. Here are some recipes for making Agar based mediums for mycelium cultures.

Enlarge

Petridishes_mycelium (1 van 2) - Potato Dextrose Agar Medium recepie

The first phase of mushroom cultivation is the isolation of a pure culture of mycelium, for growing this you need a medium that usually goes in petri dishes. Here the medium will be based on jelly agar, a seaweed derived compound which gelatinises water. The medium is basically food that feeds the mycelium while it is growing. It is possible to make different types of medium with agar with very simple and quick preparation, in this article I will describe some of the more common recipes.

But first of all: why is it useful to have mycelium in a petri dish?

- it is pure mycelium, so you don't have residuals when transferring it to other mediums or substrates
- you can store different species and samples in a small space
- it is easy to cut a little piece to start working with it
- you can save some copies of mycelium to keep the strain safely while you experiment with it (so if you fail in something, like for example if all your tests/jars get contaminated, you can easily restart cultivate it from a new piece of mycelium) cut from the original petri dish.

As I said the main medium recipes have Agar Agar as the principal ingredient, a polysaccharide gelling substance derived from marine red algae. Agar is usually mixed with others nutrients in order to create the perfect food for mycelium!

There are thousands of different recipes but the one we use in Mediamatic is Potato Dextrose Agar ( abbreviation PDA), is also the one I tried first so I can report here the recipe I used.

Potato Dextrose Agar (abbreviation PDA)
400 ml of potato broth made with 100g of potatoes
10g. agar agar
10g. unrefined cane sugar
water enough to fill up to 500 ml total
(With this recipe you'll make about 500ml of media, enough for 20 petri dishes, size 9 cm diameter)

- Boil potatoes in water with or without peel until the potatoes are cooked. Drain the potatoes, filter and reserve the liquid (and eat the potatoes preferably!)
- Mix the potato broth with agar powder and sugar in a bottle and add water until you reach 500ml.
- Put the bottle with media in a pressure cooker and sterilize it (leave the cap a bit open for the pressure to release)
- Once the pressure is high: cook for 20 minutes and let it cool down in the pan until you can comfortably hold the bottle
- Now you are ready to pour the media into the petri dish, but remember that when preparing your petri dishes you must work under the most sterile conditions you are able to achieve, so if you can get access to a lab, use a cabinal flow or alternatively, built a DIY glove box! Put the petri dish and the PDA bottle in a clean lab (or in a glove box) and before it solidifies pour it into the petri dishes and let it cool down completely.

We use disposable plastic petri dishes for general culturing, these non-reusable dishes are pre-sterilized, as an alternative is possible to use Glass petri dishes which can be heat sterilized and are reusable.

As the media cools down, it will harden into a gel, and eventually the petri dishes can be turned upside down (the media will remain in place), this is useful to avoid condensation to fill the cap of your petri dish. If petri dishes will not be used immediately, they may be stored in a bag in the refrigerator for a few weeks. If stored, be sure to check for mold or bacterial contamination before use.

According to Paul Stamets' book "Growing Gourmet and Medicinal Mushrooms" I report here some other popular recipes:

Malt Extract, Yeast Agar (abbreviation MYA, MYPA with peptone)
1 liter water
20g. agar agar
20g. barely malt sugar
2 grams yeast
1g. peptone (optional, soybean derived)

Oatmeal, Malt, Yeast Enriched Agar (OMYA, rich medium which doesn't need to be filtered although some prefer to do so)
1 liter water
20 gr. agar agar
80gr. instant oatmeal
10 gr. malt sugar
2 grams yeast

Dog Food Agar (DFA)
1 liter water
20 gr. agar agar
20gr. dry dog food

Cornmeal, Yeast, Glucose Agar (CMYA, this medium is no nutritious as the others descibed and is often used by mycological laboratories for storing coltures)
1 liter water
20 gr. agar agar
10 gr. cornmeal
5 gr. malt or glucose
1 grams yeast

Some of these recipe (especially the dog food one) may require a filtering step before sterilization to make the medium more smooth and see more easier what's going on (above all spot infections!).

Some mushroom growers say it helps to switch media to keep your strain more stable when you replicate it (when the mycelium colonize the whole agar in the petri dish is time to transfer a piece of it in a new agar petri dish for keep the strain strong).

I report also a useful summary of media commonly used with the description of their specific properties found on http://www.shroomery.org/9468/Culture-Media-for-Fungi

Water Agar (WA)--use for isolating fungi from surface-sterilized substrates.
Antibiotic Agar (AA)--use for isolating fungi from substrates not readily surface-sterilized, or to clean up a culture contaminated with bacteria.
Acidified Cornmeal Agar (ACMA)--use for isolating fungi from substrates that are likely to be contaminated with bacteria. Not a substitute for AA, but the acidity inhibits bacteria and the medium supports the growth of a wide range of fungi.
Cornmeal Agar (CMA)--use for growing a wide range of fungi, particularly members of the Fungi imperfecti; provides a good balance of mycelial growth and sporulation.
Potato Carrot Agar (PCA)--considered a relatively weak medium somewhat comparable to CMA, good for some Fungi imperfecti.
Malt agar (MA)--lacks peptone, and is useful for culturing many Ascomycota; sporulation in some species is inhibited by peptone.
Malt Extract Agar (MEA)--a good growth medium for soil fungi, fungi isolated from wood, basidiomycetes, etc. An all-purpose type of medium.
Potato Dextrose Agar (PDA)--a relatively rich medium for growing a wide range of fungi.
Potato Dextrose-Yeast Extract Agar (PDYA)---good for growing cultures derived from mushrooms.