clean clones

Jim King soma at fox.nstn.ca
Sat Mar 25 16:51:26 EST 1995


In article <3l1e3j$8e0 at bud.peinet.pe.ca>, stewart at bud.peinet.pe.ca (Paul Stewart) says:
>
>I have been establishing cultures of shiitake and oysters from grocery 
>stores to practice my clean techniques, establish my home lab, and 
>prepare for the real thing from wild specimens when things thaw out and 
>start sprouting. After exhausting all other sources of contamination, I 
>am left with the contaminants brought home from the store. (If all the 
>other produce I buy is this dirty it's no wonder my fridge looks like a 
>cool composter half the time...why do they call it a "crisper" :-) )
>
>I have tried wiping the outside of the mushrooms with ethanol (70%), but 
>still get bacteria in my cultures. I keep cultures at 25-26 C. I have 
>done lots of plant tissue culture and so tried rinsing the shrooms in 70% 
>isopropyl alcohol for 30 sec, followed by 2% bleach + 0.2% dish detergent 
>for 10 min, with shaking, then three rinses in sterile water.  The 
>oysters were glassy and translucent throughout. I squeezed out most of 
>the water, but they still looked pretty wimpy. The shiitake, being more 
>dense, held up better. Two days later, the cultures (on PDA + peptone and 
>wood) already show bacterial colonies.
>
>The question is, is there a standard disinfection procedure that will 
>clean into the gills, while still leaving viable tissue. I always take 
>tissue from the inside, just above where the stem meets the cap, 
>especially for oysters, whose cap is too thin to excise from. I'd like to 
>solve this problem before I need to clone a valuable specimen. Never had 
>this problem with chanterelle or boletus...the fungi carried into culture 
>with these species seem to be beneficial if not symbiotic.
>
>Is contamination from cloned storebought a problem others have 
>encountered and solved? (frustration mounts with each passing week!)
>
>BTW...while I'm going on and on...I posted a question recently regarding 
>using a horizontal oil drum with a hatch cut in the side as a rolling 
>spawn generator. Any comments yet on this?
>
>                        Thanx...Paul Stewart
>                        <stewart at bud.peinet.pe.ca>


Try sandwiching the contaminated culture between two layers of agar 
medium. Place the sterilized agar out of a non-innoculated petrie 
dish directly on top of your  contaminated culture. The mycelium 
should grow up thru the second layer cleaning itself by friction.
Do this as soon as you see the contaminant. I learned this from Paul
Stamets at one of his workshops. His books " Growing Gourmet and 
Medicinal Mushrooms " and " The Mushroom Cultivator " are a must for
any sincere amateur or professional mushroom grower. They cover all
aspects from scratch to finish. His company is FUNGI PERFECTI ...
e-mail:MycoMedia at aol.com Tel:360-426-9292/800-780-9126 Fax:360-426-9377

Enjoy, Somananda < soma at fox.nstn.ca >



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