Saving Strains with Agar

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By Nan (Nanook) on Tuesday, October 23, 2001 - 05:40 pm: The Nook

The Traditional Strain Saving Tek
You will need the following:

Pressure Cooker/Canner
A glass bottle with a narrow neck (like a Mrs. Butterworth bottle, or an autoclavable flask)
Light Powdered malt (Home Brew store)
Agar Agar (health food store)
Inoculation loop
A supply of 20 mm x 100 mm test tubes
Petri Dish(es)

First things first. The main workings of this have been made possible by the book, The Mushroom Cultivator, by Paul Stamets. I highly recommend purchasing it (money VERY well spent) from Fungi Perfecti. Now let us save strains, shall we?

First Mix your dry ingredients together (Yeast, Malt Powder, and Agar). Add aproximately 50 grams of this mixture to 1 liter of water (the liter is already in your glass bottle). Make sure that the bottle doesn't get filled within 3/4 of its capacity. Shake the bottle (with the water and dry ingredients within) to mix the water and dry ingredients evenly. Plug the hole of the bottle with a cotton ball, and then cover the top with aluminum foil. Then place the bottle in the pressure cooker/canner and sterilize at 15 psi for 30 minutes (follow the instructions included with the cooker). Let the pressure go back down to 0 before opening the cooker/canner. Remove the bottle from the cooker/canner, and the tin foil. When ready to pour, unplug the cotton ball and pour the liquid into the petri dish(es) and test tubes. Fill the test tubes only 1/3 of the way up, and let the test tubes rest on something (piece of wood, book, etc) at a 15 to 30 degree angle. Also, cover the petri dishes with their lids. Let the agar solidify in the test tubes and petri dishes. When cool, and solidifies, rub the inoculation loop across a spore print, and then across the surface of the agar in the petri dish. Let the spores germinate, which will take several days. Do NOT let the mycelium cover the entire petri. Flame sterilize (lighter, or other alchohal flame are good for this) your scalpel. Cut a small square of mycelium from the petri (small enough to fit into the test tube). Then transfer the square to one of the test tubes. Let the mycelium grow across the top of the agar inside of the test tube. This will only take a couple days. Do not let the mycelium go to far deep into the agar (ie, no more than a cm or so). Then plug the top of the test tube with a cotton ball, and screw the cap on (if your test tubes do not have caps use tin foil to cover them). Mark the strain and date on the test tube, and refrigerate until needed! Take the test tube out every couple of months, and transfer (with a sterile scalpel) some mycelium to a petri, then start the whole process over if necessary. If not, put the test tube back in the fridge! And remember... DON'T BE AFRAID OF AGAR!!! It will speed up your colonization times, and make your life a lot easier in the long run!!!

Have fun!

Agar Culture Info : Shroom Glossary