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OfflineSclerotiogenesis
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Registered: 06/20/15
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Optimized agar for sclerotiogenesis
    #21833745 - 06/20/15 08:09 PM (8 years, 7 months ago)

I'm attempting to produce highly sclerotiogenic strains of Psilocybe armandii and Psilocybe semilanceata, two species known to naturally produce small sclerotia. All the most high yielding sclerotium producers are strains that resulted from artificial selection for that phenotype e.g., ATL#7 was selected by workman for its large sclerotia and the tamapnensis strain in circulation was selected by Pollock for its large sclerotia, thus I think it's unfair to dismiss a species for naturally producing small stones until some efforts have been made to see if the size/number can be increased.

I am going to be screening these species for sclerotia production on agar. My plan is to inoculate at least ten plates for each species with a dilute spore solution and look for natural sclerotium production as strains that do this generally perform excellently when introduced to grain, but my concern is that no sclerotia will naturally form on the agar.

So here is my question: how can an agar formula be developed that's optimized for inducing sclerotiogeneis?

It's clear that sclerotia favor rye grain or rye seeds, so I wonder if I should make some sort of rye grain extract based agar. If I were to do this would the extract ideally be an aqueous rye grain extract or something made with an organic solvent such as MeOH? Also I wonder if the addition of white quartz sand to the agar would provide a sort of nidus or nucleation point for the sclerotia.

Any thoughts or feedback would be appreciated.


Edited by Sclerotiogenesis (06/20/15 08:32 PM)


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Invisiblemush madness
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Re: Optimized agar for sclerotiogenesis [Re: Sclerotiogenesis]
    #21833801 - 06/20/15 08:24 PM (8 years, 7 months ago)

You want 4.5 % malt and add 1 gram instant coffee per 1000 ml

That is what I have found to work the best for me anyways


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OfflineSclerotiogenesis
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Re: Optimized agar for sclerotiogenesis [Re: mush madness]
    #21841806 - 06/22/15 02:13 PM (8 years, 7 months ago)

Thanks for the tip. Has anyone attempted to make a rye grain or seed extract as an agar additive? It's unclear to me whether these substrates are ideal for sclerotia production due to their physical characteristics (hard and allowing air penetration), chemical composition, or both.


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Offlinematsc
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Re: Optimized agar for sclerotiogenesis [Re: Sclerotiogenesis]
    #21842047 - 06/22/15 03:18 PM (8 years, 7 months ago)

In nature sclerotia formation is mostly a function of local conditions, rather than specific substrate. Changes in moisture, temperature, oxygen content, nutrient levels, etc. Its a survival strategy the fungus implements when it senses hard times are ahead.

There are several functions the sclerotia themselves serve, depending on the species. Some are water storage reserves, others are the primary inoculum, and quite often theyre a means to survive brush fires. This last one is the current leading hypothesis for the Psilocybe sclerotia formers. Its likely a combination of nutrient depletion and dropping moisture content, such as a mushroom in a grassy field might experience at the end of spring as the summer fire season approaches that triggers the process (in general terms, specifics are fuzzy here).

Honestly, if you want to select for the largest sclerotia, you should clone from large sclerotia. Pick out the biggest one you can lay your hands on, take a sterile sample and plate it, and repeat till you end up with what you want. If youre working from a spore sample originally, you will have mixed genetics to start, but you can clone to single isolates within a generation or two.


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OfflineSclerotiogenesis
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Re: Optimized agar for sclerotiogenesis [Re: matsc]
    #21874401 - 06/29/15 04:58 PM (8 years, 6 months ago)

Quote:

matsc said:Honestly, if you want to select for the largest sclerotia, you should clone from large sclerotia. Pick out the biggest one you can lay your hands on, take a sterile sample and plate it, and repeat till you end up with what you want. If youre working from a spore sample originally, you will have mixed genetics to start, but you can clone to single isolates within a generation or two.




As stated in the OP, I am working with rare species that are poor sclerotia producers. I think under optimum conditions both species will produce small sclerotia after several months but I would prefer to pre-select on agar before inoculating grains.


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Offlinematsc
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Re: Optimized agar for sclerotiogenesis [Re: Sclerotiogenesis]
    #21874492 - 06/29/15 05:16 PM (8 years, 6 months ago)

Ah, my apologies, I read the sentence about ATL7 being selected for large sclerotia and my brain somehow decided you were attempting the same. 

That said, I still think it would be a combination of physical and chemical signals that would induce sclerotia formation. Use half or quarter strength agar (in terms of nutrients, not agar percentage). Let the agar dry out a bit over time. Warm it up a bit. Try to trip as many of the signal pathways as you can that would tell the fungus "hey, summer is coming, you should head underground and prep for fire damage".


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