This guy is a local mushroom grower (oysters and the like) who has a PhD in Biochem and a Masters in molecular Bio. Here is his way to reduce contam in agar.
I am clipping the text from the following source "Growing mushrooms the easy way, Home Mushroom cultivation with Hydrogen Peroxide Volume 1" R.Rush Wayne PhD
To be on the safe side with my plate cultures, I use the lowest concentration of peroxide that I have found effective in agar medium, which is about 0.018%, or 6 mls per liter of medium. (You can add twice this much without visible harm to the mycelium of the species I have grown, but note that very slow-growing species such as Stropharia may be more sensitive to peroxide exposure.
1) I add all the ingredients to a jar with the desired amount of water. The jar should hold about twice the volume you will actually use, to keep the agar from boiling over when it cooks. 2) I adjust the pH with a little baking soda (my water is acidic, but you could use vinegar if yours is alkaline. Also, see my "Note on Measuring pH of Substrate" below in the section on preparing bulk substrate). 3) I then use my ordinary kitchen pressure cooker to melt and sterilize the medium. (I use tap water and have not had any problems with it. In fact, when I grew mushroom mycelium on medium prepared with distilled water, growth was noticeably slower). I put lids loosely in place and pressure cook at 15 psi for no more than 10 minutes, allowing an initial ten minutes of steaming to melt the agar before putting on the pressure regulator. (If you are using ready-mixed MYA medium, the instructions may say to pressure cook for much longer times, for example, 45 minutes. Don't do it! 20 minutes is the most you?ll need, and any longer will produce carmelization products in the medium that are harmful to the mycelium). I also sterilize a set of petri dishes along with my medium, placing the dishes in a large tomato can covered with aluminum foil (I use plastic reusable petri dishes, and a liter of medium fills up about 30 plates). 4) When the cooking is finished, I gently slide the cooker off the burner and allow the pressure to drop. I carefully remove the jar containing the medium and let it cool in the open air on my counter top. There is no need to avoid entry of unsterilized air, assuming there is not a great deal of heavy dust, since the peroxide will kill the airborne contaminants when it is added. 5) When I can handle the jar quite comfortably, I usually put the jar of agar in a pot of warm water for the last part of the cooling process, since the agar is close to solidifying at this temperature. 6) Then I add my peroxide solution with a pasteurized pipette and quickly mix the peroxide into the medium by moving the jar with a circular motion, reversing directions a couple of times (but doing my best to avoid making a lot of bubbles, which will end up on the surface of the agar). 7) Once I've added the peroxide, I go straight to my petri dishes, which I have set out on a clean counter, and I free-pour the medium into the dishes, closing the lids as I finish.
-------------------- Mycology is a lot like quantum mechanics in that we don?t have causal relationships like in Newtonian physics, only probabilities of various outcomes.<=== Misapropriated from Mycofile
|