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InvisibleZombiWurm
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Dehydrating mycelium
    #26354352 - 11/30/19 05:47 PM (4 years, 1 month ago)

Is it possible to preserve a liquid culture by dehydrating mycelium. Like maybe put the culture into a centrifuge to separate from the water. Would drying the mycelium work to preserve it. From what i understand, mycelium should go dormant with a lack of oxygen and water.


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InvisiblemushboyMDiscord
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Re: Dehydrating mycelium [Re: ZombiWurm]
    #26354397 - 11/30/19 06:17 PM (4 years, 1 month ago)

why not put an agar plate on ice? Making the lc to dehydrate later on is kinda pointless imo:shrug:

If I understand the process correctly. drying myc will denature certain enzymes rendering the dehydrated culture 'dead'


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Offlinecurious.psychonaut
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Re: Dehydrating mycelium [Re: ZombiWurm]
    #26354409 - 11/30/19 06:27 PM (4 years, 1 month ago)

Room-temp dried stipe tissue (no spores), which is just specialized myc, revives on agar per search results. (Using a high-temp dehydrator would indeed kill/denature proteins, starting as low as 41C.)

So there's a good chance it might work for LC as well? If it does, I guess it would probably preserve better than LC at room temp (but maybe worse than LC at 2C?--no idea). If preservation of a myc culture is the goal and you haven't looked at slants+wooden sticks yet, check them out.


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InvisiblemushboyMDiscord
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Re: Dehydrating mycelium [Re: curious.psychonaut]
    #26354419 - 11/30/19 06:31 PM (4 years, 1 month ago)

Quote:

curious.psychonaut said:
Room-temp dried stipe tissue




That's not dry:oldman:


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OfflineShroomerquest
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Re: Dehydrating mycelium [Re: mushboy]
    #26354496 - 11/30/19 07:12 PM (4 years, 1 month ago)

Quote:

mushboy said:
why not put an agar plate on ice? Making the lc to dehydrate later on is kinda pointless imo:shrug:

If I understand the process correctly. drying myc will denature certain enzymes rendering the dehydrated culture 'dead'





Can you freeze mycelium...really???!!!

If I have misunderstood, what do you mean by put agar dish on ice. Any other way I’m thinking the ice would melt and would need maintenance so why not opt for a cold refrigerator if that’s the case....

...and if I do follow correctly, how far can the freezing process be took?


I love this place, just discussing, sharing and learning from each other is priceless!!!

Where else would you get it...?:cheers:




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InvisiblemushboyMDiscord
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Re: Dehydrating mycelium [Re: Shroomerquest]
    #26354513 - 11/30/19 07:26 PM (4 years, 1 month ago)

Put on ice = slant and put in fridge.


Sorry should of been clearer
:towelie:


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OfflineShroomerquest
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Re: Dehydrating mycelium [Re: mushboy]
    #26354548 - 11/30/19 07:50 PM (4 years, 1 month ago)

Quote:

mushboy said:
Put on ice = slant and put in fridge.


Sorry should of been clearer
:towelie:




So just to be clear, excuse me if I sound rude

is ice involved here or is it some loose term for putting a slant into cold conditions

Can mycelium be froze yes or no would settle that point and if so or not so, can you elaborate please

Cheers dude


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InvisiblemushboyMDiscord
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Re: Dehydrating mycelium [Re: Shroomerquest]
    #26354577 - 11/30/19 08:14 PM (4 years, 1 month ago)

..loose term
:thatsayes:

could you freeze myc and revive it? probably not:justdontknow:


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Offlinecurious.psychonaut
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Re: Dehydrating mycelium [Re: mushboy]
    #26354601 - 11/30/19 08:32 PM (4 years, 1 month ago)

newtomyc is currently trying to recover frozen myc (though IIRC it's not clear if all was indeed frozen). It's not pretty.

Basically, the tiny ice crystals that start to form when freezing rupture cell membranes which kills them. So it's not a way to store myc (or even fruits for consumption, for the same reason: they lose their structure).


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OfflineShroomerquest
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Re: Dehydrating mycelium [Re: mushboy]
    #26354612 - 11/30/19 08:40 PM (4 years, 1 month ago)

Quote:

mushboy said:
..loose term
:thatsayes:

could you freeze myc and revive it? probably not:justdontknow:





Thanks mushboy


Quote:

curious.psychonaut said:
newtomyc is currently trying to recover frozen myc (though IIRC it's not clear if all was indeed frozen). It's not pretty.

Basically, the tiny ice crystals that start to form when freezing rupture cell membranes which kills them. So it's not a way to store myc (or even fruits for consumption, for the same reason: they lose their structure).




This was my stand on the matter

Kinda got hopeful and excited there for a second :rofl:


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Re: Dehydrating mycelium [Re: Shroomerquest]
    #26354621 - 11/30/19 08:44 PM (4 years, 1 month ago)

So what about dried myc

Reminds me of the story I heard of soma and how it fits with the wooden drinking utensil was harbouring mushroom mycelium that when soaked, grew and that it could’ve consumed


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InvisibleZombiWurm
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Re: Dehydrating mycelium [Re: mushboy]
    #26355277 - 12/01/19 10:00 AM (4 years, 1 month ago)

Thanks for the response. What enzymes would become denatured? As for the freezing, I assume mushroom mycelium could be frozen with cryogenic (adding antifreeze as the water is removed) I wonder if it would survive the whole process and thaw?


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OfflineeLeSDenes
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Re: Dehydrating mycelium [Re: ZombiWurm]
    #26355323 - 12/01/19 10:30 AM (4 years, 1 month ago)

i don't really get why would you go through all the pain of drying mycelium when you could just make culture slants. I just removed a 5 year old galindoi culture from the fridge which I haven't touched since I made them. I transferred some to agar plates and they look perfect.


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InvisibleZombiWurm
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Re: Dehydrating mycelium [Re: eLeSDenes]
    #26355342 - 12/01/19 10:46 AM (4 years, 1 month ago)

Longer term storage. Under the right conditions, biological material can survive for hundreds of years or more. It started as just a thought when I was trying to sleep.


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Offlinecurious.psychonaut
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Re: Dehydrating mycelium [Re: ZombiWurm]
    #26356330 - 12/01/19 09:49 PM (4 years, 1 month ago)

Found this interesting and researched a bit.

[To recap, for the hobbyist looking for methods without specialized equipment that are proven and well-known in the shroomery: Slants at 2C store a culture for years (perhaps decades, if refreshed regularly). Dry prints store a variety for many years (10+ in ideal conditions). A print pool refreshed via fruiting+printing = indefinite storage.]

Survival for years of fungi in dry wood has been documented since the 1920s. (We also put wood in slants.) A method for storing under oil was developed in the 1940s (sci-hub).

For longer term storage of revivable hyphae, the gold standard, developed in the 1960s, seems to be liquid nitrogen refrigeration in glycerol water (10% vol.) which works for an indefinite amount of time (as long as temp is kept below -130C).

As can be seen in the 60s paper, Basidiomycetes (the large division of higher fungi to which cubes belong) were thought to be incapable of surviving freeze-drying at the time. But newer methods were developed in the 90s that were demonstrated to partially work (sci-hub) for Basidiomycetes and also specifically on a member of Agaricales (the gilled shrooms, the smaller group incl. cubes). The trick was to add 5% trehalose to the growth medium (instead of later on as preservant). However, the revived hyphae did not produce fruiting bodies :sad:.

Quote:

The survival rates of the two Basidiomycetes studied were found to be lower than those of the Ascomycetes. Revived colonies of Schizophyllum commune [the gilled shroom included in the study], incubated in MP medium [malt-peptone agar without trehalose], were extremely poor and slow-growing; basidia [the spore-producing structure at the gills] were not produced. When freeze-dried after incubation in MPtreh [malt-peptone agar with trehalose], a higher growth rate was obtained but colonies remained atypical as no aerial mycelium and no basidiomata [actual shrooms] were produced




The intro of the above study provides an overview of current methods:

Quote:

Methods of long-term preservation in which metabolism is inhibited during storage are preferable to maintenance on agar slants. The last mentioned method is laborious because regular transfers onto fresh media are required. In addition, repeatedly subcultured strains may undergo mutations.

Current methods for long-term preservation are freeze-drying and preservation at temperatures below -130C. Cryopreservation methods give high survival rates and are universally applicable. However, some disadvantages are encountered. Methods using nitrogen are rather expensive. Regular supply of liquid nitrogen is not always guaranteed. [...]

Freeze-drying is a good alternative. Ampoules can be stored easily in dense packing without any special requirements. Cultures need not be revived on agar slants prior to dispatch. The product is light, inactive and dry, enabling easy distribution by mail.

However, so far only conidia or spores can be freeze-dried successfully. Most attempts to revitalize dehydrated hyphae have failed, except [...]

Previous studies have shown that the survival rate of frozen fungal cells increases considerably when cells are cooled at a rate of -1C/min. Several authors have stressed the beneficial effect of trehalose. Addition of peptides to the resuscitation medium is aimed to improve recovery. After optimization of these parameters, hyphae of a test series of Ascomycetes and Basidiomycetes were freeze-dried. Trehalose was added to the growth medium rather than to the protectant to allow absorption by the cell. Absorption would result in protection of cellular membranes as well as membranes of organelles




I was unable to find studies examining revival of air-dried/centrifuged hyphae, so I suppose it's deemed a less viable option than freeze-drying. It's interesting to note that many other fungi can survive freeze-drying or even natural freezing for at least 140,000 years (sci-hub). So maybe we just need to find the right procedure and medium/protectants...


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Edited by curious.psychonaut (12/01/19 10:17 PM)


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InvisibleZombiWurm
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Re: Dehydrating mycelium [Re: curious.psychonaut]
    #26356972 - 12/02/19 10:27 AM (4 years, 1 month ago)

That is very interesting. Glycerin is used to in humans for cryogenics I believe. I thought freeze drying things destroyed cell structure. I've had freeze dried cubes before and they made me itch really bad. I'm still not sure if that means anything or not but I've never itched from cubes before. They looked really awesome though.

So basically I can store spores longer than I will live but not mycelium, as of right now. I'm okay with that lol. How do I store spores to be reliably revived over a hundred years from now? Just freeze dry them and store in liquid nitrogen?


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OfflineThe Mycologist
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Re: Dehydrating mycelium [Re: ZombiWurm]
    #26356980 - 12/02/19 10:31 AM (4 years, 1 month ago)

Best way to store genetics for long term is to put them in a agar slant with a tiny piece of wood, and leave them in a lunchbox in the fridge.

This technique can keep a culture for years.


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Offlinecurious.psychonaut
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Re: Dehydrating mycelium [Re: ZombiWurm]
    #26357014 - 12/02/19 11:02 AM (4 years, 1 month ago)

Quote:

ZombiWurm said:
How do I store spores to be reliably revived over a hundred years from now? Just freeze dry them and store in liquid nitrogen?




Liquid nitrogen seems to be the best for both spores and myc (the freezing procedure is different from freeze-drying though).


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InvisibleZombiWurm
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Re: Dehydrating mycelium [Re: curious.psychonaut]
    #26357023 - 12/02/19 11:11 AM (4 years, 1 month ago)

Okay. I have some wild prints and 1 cube print in my freezer right now and went to transfer Psilocybe Cyanescens spores to agar a few weeks ago and the only thing that grew was green.


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