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OfflineMonkeyJesusFresco
am i suspended in agar?
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Wild Tissue Culturing
    #21696589 - 05/18/15 10:58 PM (8 years, 8 months ago)

1. So I've been fooling around with these little agar containers.
2. I have a tendency to collect wild fruit-bodies and then culture them.

Long story short:

I read something somewhere, in a book by Paul Stamets and I came up with this:



So, what you see there is a 3.5 fl. oz. polypropylene container, with about 2 tablespoons of light-malt-extract-agar and a coffee-stir-stick cut down to size, sticking up.

I came across some wild fruit-bodies that I wanted to culture, unfortunately they had been out there for a while...and they sat in the refrigerator for some time, as I simply never got around to fooling with them and... needless to say that, in the end, clean inner-stipe tissue was not gonna be an option for culturing :shrug:

So what I did was:

-Chop up a stem

-Soak stem pieces in a diluted dishsoapy-water for 15-20 mins

-in a still air box, with tweezers, I picked each stem piece out of the soapy water, and rinsed it off in a 1:10 bleach:water solution for an unknown amount of time (I did a double-blind study on myself, a.k.a. I didn't take lab notes)

Anyways, in the end I had 6 of the containers that you see pictured above, each with a "washed" piece of mushroom stem laying on or near the intersection of the stick&agar. Each piece was washed off for different unknown times in the bleach solution; (no longer than 1 minute)

So, the idea was to "leave all the contaimination behind" on the agar as the "wood-loving" mycelium from the mushroom stipe, crawled up the stick, in the hopes that, at a later date, I could come back and retrieve a clean culture. (From what I recall, Paul Stamets version of this used whole pieces of straw)

In conclusion:

- I desperately need to get my hands on some Gentamycin Sulfate, as 5 out of the 6 containers that I made were over run with bacteria, not to mention MOLD!

- Oddly enough, one of the containers grew mycelium and no bacteria or mold (the "pride&joy" pictured above), at the moment I'm contributing this "low success rate" to the varied wash times in the bleach-water bath, and possibly the small amount of soap used in the soap bath.

And finaly:

Does anyone have anything to contribute to this topic? Experience? Ideas? Anything???

I can't remember where I got the idea to run the mycelium up a stick like that, and last time I mentioned something like it (on a different website) I was very heavily ridiculed for it :confused:

So hopefully this post will open a line of discussion and informative input.
:laugh:


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InvisiblebodhisattaMDiscordReddit
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Re: Wild Tissue Culturing [Re: MonkeyJesusFresco]
    #21697186 - 05/19/15 05:15 AM (8 years, 8 months ago)

Skip the soap and bleach use those and anti biotic agar as a last resort try it the normal way first. Inner tissue biopsy. Then the mycelium is less injured will recover faster and you have a fighting chance of it over running bacteria and if your sterile technique is good you should get 0 mold.


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OfflineMonkeyJesusFresco
am i suspended in agar?
Male


Registered: 10/09/12
Posts: 3,306
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Last seen: 4 hours, 11 minutes
Re: Wild Tissue Culturing [Re: bodhisatta]
    #21697710 - 05/19/15 09:53 AM (8 years, 8 months ago)

Yeah, I had samples collected from two different patches; the first patch/sample, the mushroom had been plucked from the ground, the entire fruit body was intact, so inner-stipe tissue was clean and there was no problem culturing that; the second batch was old/weathered, had been cut and neglected in the fridge for some time, so I rigged this up, basically washing cut-up stipe pieces off; either way, I got clean cultures from both patches :cool:

and yes, all that non-sense should be left as last resort :super:
especially the "washing", took about a week-and-a-half to two to recover :shrug:

edit: also, the circumstances I'm working under, resources are TIGHT. no petris dishes, much less antibacterial agar :frown:


Edited by MonkeyJesusFresco (05/19/15 09:56 AM)


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