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 x7x_x7x
urban farmer
Registered: 05/05/07
Posts: 668
Last seen: 1 hour, 3 minutes
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 experimental tek for obtain micelium from dirty spore prints
#8362290 - 05/04/08 06:22 PM (5 months, 1 hour ago) |
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the idea is:
make a spore syringe from a wild collected specimen.
add 1 - 2 cc to an antibiotic agar
let rest 24 -48 hs
transfer the liquid to another petri dish with antibiotic agar
let rest 24 -48 hs and so
the concept is: molds are fast colonizers than basidiomycetes. the spores of fungical contams grow in the first petri dishes, the last transfers contain only the spores of slower basidiomycetes.
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transhumanism + primitivism=magic mushrooms
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 fastfred
Registered: 05/17/04
Posts: 4,326
Loc: Dark side of the moon
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 Re: experimental tek for obtain mycelium from dirty spore prints [Re: x7x_x7x]
#8364210 - 05/05/08 07:58 AM (4 months, 30 days ago) |
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That's pretty standard technique. You're going to want to use a lot less than 1-2cc on a plate though. The idea is to dilute the mixture to the point that you only get a handful (or less) colonies per plate. Then you can easily isolate what you're looking for.
So you're going to want to use a cc or less of pretty dilute spore water. A more used method is to streak your solution or dry spores on a dish. If you just put solution in your dish you will likely have problems with yeast contamination. If you have solution sloshing around in your dish any yeast in the mix is going to be distributed over the entire dish, contaminating it. Your best bet is using a small amount of dilute solution or using dry spores. I'd use a pretty soft agar mix or you might have problems getting dry spores to germinate.
The easiest way is to just tap a cap over a dish a couple times and then isolate.
-FF
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x7x_x7x
urban farmer
Registered: 05/05/07
Posts: 668
Last seen: 1 hour, 3 minutes
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Re: experimental tek for obtain mycelium from dirty spore prints [Re: fastfred]
#8368023 - 05/06/08 07:39 AM (4 months, 29 days ago) |
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i'm using this tek and the contam load is lower in each transfer; will be interestig if other people reproduce the experiment
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transhumanism + primitivism=magic mushrooms
SuctoSporeŽ
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MycoAu
Registered: 07/18/07
Posts: 470
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Re: experimental tek for obtain mycelium from dirty spore prints [Re: x7x_x7x]
#8368662 - 05/06/08 12:18 PM (4 months, 29 days ago) |
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FF, you mentioned tapping a cap above an open plate for beginning the isolation procedure. Done in a hood? Or will it matter because of the contaminate load from the mushroom itself? Plus, how do you ensure that any of the spores actually make contact if it is performed in the hood? (afraid that they'd be carried away in the flow- as one might expect in a hood)
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solumvita
Registered: 02/12/08
Posts: 34
Last seen: 2 days, 7 hours
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Re: experimental tek for obtain mycelium from dirty spore prints [Re: MycoAu]
#8368688 - 05/06/08 12:29 PM (4 months, 29 days ago) |
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MycoAu, the simplest would be to hold the lid of the petri dish in front of the flow so that the cap is shielded from the air flow.
Also I agree with FF in that rather use dry spores, my problem is normally bacteria contaminate especially as soon as water is added.
ciao
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lipa
Registered: 07/24/07
Posts: 502
Last seen: 6 hours, 51 minutes
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Re: experimental tek for obtain micelium from dirty spore prints [Re: x7x_x7x]
#8368751 - 05/06/08 12:54 PM (4 months, 29 days ago) |
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I usually take the wild mushroom cap and place it on top of a aseptically cleaned regular or wide mouth jar band over paper or foil and swab the cap with light ball of alcohol. I cover it and let it drop spores overnight to drop the dirty spores. Then I transfer to another clean lid band and repeat to obtain newer cleaner ones. The second set is usually pretty darn clean. In fact, Contaminants seem kinda rare after switching to this method. I think most of the contaminants usually come from being in contact with the print medium itself.
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lipa
Registered: 07/24/07
Posts: 502
Last seen: 6 hours, 51 minutes
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Re: experimental tek for obtain mycelium from dirty spore prints [Re: solumvita]
#8368757 - 05/06/08 12:58 PM (4 months, 29 days ago) |
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Quote:
solumvita said:
MycoAu, the simplest would be to hold the lid of the petri dish in front of the flow so that the cap is shielded from the air flow.
Thats a very bad habit! Working in front of the hood with spores. 
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MycoAu
Registered: 07/18/07
Posts: 470
Last seen: 7 hours, 13 minutes
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Re: experimental tek for obtain mycelium from dirty spore prints [Re: lipa]
#8370231 - 05/06/08 07:02 PM (4 months, 29 days ago) |
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I've always just created a spore print- didn't care whether it was dirty or not. Then, scrape into sterile water, inoculate small amounts of wbs and transfer colonized grain after it has grown UP into the upper grain. This obviously doesn't solve mold problems, but I've never had a problem with bacteria using this particular method.
I'm always curious about better, easier methods.
Oh, and don't forget the power of gentamicin, chloramphenicol, etc.
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solumvita
Registered: 02/12/08
Posts: 34
Last seen: 2 days, 7 hours
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Re: experimental tek for obtain mycelium from dirty spore prints [Re: lipa]
#8377161 - 05/08/08 11:39 AM (4 months, 27 days ago) |
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Sorry lipa, I just can't win  I must say i never thought of it, but i do have two hoods and a progression of labs, the first being my dirty one and then the sterile one. Certain work is never performed in clean lab and vice versa.
i take your point though. Where do you transfer spores, do you use a glove box?
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lipa
Registered: 07/24/07
Posts: 502
Last seen: 6 hours, 51 minutes
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Re: experimental tek for obtain mycelium from dirty spore prints [Re: solumvita]
#8377203 - 05/08/08 11:55 AM (4 months, 27 days ago) |
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In a still air box sometimes. But really I see no need for that even. Your going to have to transfer either way, so most of the time I just do it in the open. Blowing spores all over the place "in my opinion" is a very bad thing.
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lipa
Registered: 07/24/07
Posts: 502
Last seen: 6 hours, 51 minutes
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Re: experimental tek for obtain mycelium from dirty spore prints [Re: solumvita]
#8377421 - 05/08/08 12:44 PM (4 months, 27 days ago) |
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Quote:
solumvita said:
Sorry lipa, I just can't win
By the way! This is not a win or lose game. This is a helping game and there are no losers.
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BlimeyGrimey
Pyon!
Registered: 08/24/05
Posts: 872
Loc: Seattle, Wa
Last seen: 3 hours, 10 minutes
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Re: experimental tek for obtain micelium from dirty spore prints [Re: x7x_x7x]
#8381146 - 05/09/08 09:36 AM (4 months, 26 days ago) |
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Streaking spores onto agar in a zig-zap pattern then transfering good looking growths to another petri is your best bet. Adding large amounts of liquid to an agar dish is just asking for bacterial contams.
Transfering the liquid to another dish will simply carry any growing mold/bacteria to the next dish. They wont simply decide to stick to the first dish , some spores/living mold will still be in the water. Also by transfering growing mold/bacteria to another dish simply helps it grow faster since its already started the process. Doing this on antibiotic agar would simply help you transfer resistant contams from the first dish to the second.
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Confirmed:
Psilocybe stuntzii
Psilocybe cyanescens
Psilocybe guilartensis
Panaeolus olivaceus
Unknown:
Washington Psilocybe species
Washington Panaeolus species
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x7x_x7x
urban farmer
Registered: 05/05/07
Posts: 668
Last seen: 1 hour, 3 minutes
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Re: experimental tek for obtain mycelium from dirty spore prints [Re: BlimeyGrimey]
#8382707 - 05/09/08 06:22 PM (4 months, 26 days ago) |
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Quote:
BlimeyGrimey said:
Transfering the liquid to another dish will simply carry any growing mold/bacteria to the next dish. They wont simply decide to stick to the first dish , some spores/living mold will still be in the water. Also by transfering growing mold/bacteria to another dish simply helps it grow faster since its already started the process.
i try this tek and the mold amount decreases in each transfer. the first plate is a freak zoo, the next got only one or two mold colonies and one more give only basidios.
Quote:
Doing this on antibiotic agar would simply help you transfer resistant contams from the first dish to the second.
in my experience, growing spores from wild specimens in plain agar results in a bacterial disaster. genta and chlora are old and widely used antibiotics, and resistance to both are not a health hazard. bacterial development in agar treated with both atb's is very rare; as you can read in any microbiology book. if the case, simply add 70% alcohol and discard. no microorganism resist 70% alcohol.
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transhumanism + primitivism=magic mushrooms
SuctoSporeŽ
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