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Offlinesstox
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[Q] About isolating monoculture and its aging process
    #21473282 - 03/29/15 06:09 AM (9 years, 1 month ago)

Hi @ all,
I'm new to mushroom cultivation and experiencing with pc mckennaii for the first time. Therefor I'm looking for a couple of answers to questions/confirmations of thoughts which came to my mind lately.

So cultivating... I transfered mycelium from a growkit to several grain jars using a laminar flow hood. It grows pretty good. I could go on with this and transfer to the next jars, but I think in long terms it would be best to isolate a monoculture from the master mycelium.

So my 1st question is about isolating the monoculture.
After reading several tutorials by Paul Stamets and Marc Keith I think I could be able to isolate a rizomorph monoculture. I would isolate different parts of a culture and separate them from each other. After that I would do tests, which one grows best. BUT: This way I would only be able determine, which culture grows best but not the amount of psilocybin and psilocin. I guess pure mycological forums would kick me for such a question. ;-) Can I (and if, how) determine the amount for psilocybin and psilocin from the different cultures? Or shouldn't I bother about this and just care about the growth strength? Please note, I do not have a laboratory or access to special chemicals.

The second thing is more than an assumption which I am looking a confirmation or declination for. I can't use a "master" monoculture forever, can I? Even with long-term storage techniques (btw: cool guide here with distilled water ;-)). Like every cell divisioning organism, the culture can get older, lose its vitality and strength and finally dies. Is that correct? Or can it theoretically exist forever? Should I keep spore prints from time to time in case the master culture gets old to germinate a new culture?

Many thanks in advance for your thoughts.

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InvisibleATXfungi
Mycology Student


Registered: 02/28/15
Posts: 74
Re: [Q] About isolating monoculture and its aging process [Re: sstox]
    #21474208 - 03/29/15 11:58 AM (9 years, 1 month ago)

Quote:

sstox said:
Can I (and if, how) determine the amount for psilocybin and psilocin from the different cultures? Or shouldn't I bother about this and just care about the growth strength?





Measuring the amount of actives in your cubes is going to be outside your capabilities unless you have access to a university lab or private lab. You can always do a bioassay(eat some) and judge potency subjectively, but that is going to be influenced by many factors other than just the dose.

More important than isolating a strain that produces alot of alkaloids is refining your substrate and growing environment. A well prepared sub will do more for the potency than putting in all that isolate testing.

Quote:

sstox said:
I can't use a "master" monoculture forever, can I? Should I keep spore prints from time to time in case the master culture gets old to germinate a new culture?





Depends on how you choose to store the culture. If you keep it frozen with nitrogen it should last (practically) forever. Most the methods home users favor aren't as good. Store your cultures as many ways as you can and see which methods give you the best results. Keep duplicate cultures in a different location as a backup library...just in case. Definitely make spore prints. Prints are easy, resilient, take up no space, and are good to trade with.


--------------------
“We need to make books cool again. If you go home with somebody and they don't have books, don't fuck them.”-John Waters

“Insanity is relative. It depends on who has who locked in what cage.”-Ray Bradbury

"The surest way to corrupt a youth is to instruct him to hold in higher esteem those who think alike than those who think differently."-Nietzsche

"There are no passengers on spaceship earth. We are all crew."-Marshall McLuhan

Edited by ATXfungi (04/01/15 06:16 PM)

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Offlinesstox
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Re: [Q] About isolating monoculture and its aging process [Re: ATXfungi]
    #21481374 - 03/30/15 10:07 PM (9 years, 1 month ago)

Hi ATXfungi and thanks a lot for the reply. Oki, I'll isolate a the monocultures, do some potency-tests and keep the best. Boy, that's going to be an awesome spring ;-)

Of course I do not have nitrogen at home. So I'll just try to keep the best strain alive as long as possible with slants and also do spore prints as plan B.

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InvisibleATXfungi
Mycology Student


Registered: 02/28/15
Posts: 74
Re: [Q] About isolating monoculture and its aging process [Re: sstox]
    #21482301 - 03/31/15 06:27 AM (9 years, 1 month ago)

Sounds like you've got a good plan.

Don't forget about the DIH20 method you mentioned earlier. I've got some cultures that are about 5 years old using that method that I'm planning to test for viability this summer. I'll post the results.

Enjoy the isolations. For me, agar work is meditative.


--------------------
“We need to make books cool again. If you go home with somebody and they don't have books, don't fuck them.”-John Waters

“Insanity is relative. It depends on who has who locked in what cage.”-Ray Bradbury

"The surest way to corrupt a youth is to instruct him to hold in higher esteem those who think alike than those who think differently."-Nietzsche

"There are no passengers on spaceship earth. We are all crew."-Marshall McLuhan

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Offlinesstox
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Re: [Q] About isolating monoculture and its aging process [Re: ATXfungi]
    #21485767 - 03/31/15 11:05 PM (9 years, 1 month ago)

Cool, please share the results. Which technique did you use?
Practical method for long term storage of mycellium in dH2O using ependorf tubes as vials.
or
Resurrecting a Better Method for Long-Term Storage of Mushroom Cultures
Since they have different methods gaining the mycelium off the petri dish.

Did you use deionized water? The threads talked about distilled water. Would that make a difference?

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InvisibleATXfungi
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Posts: 74
Re: [Q] About isolating monoculture and its aging process [Re: sstox]
    #21488583 - 04/01/15 04:38 PM (9 years, 1 month ago)

I used this as my general guide. http://www.ncbi.nlm.nih.gov/pmc/articles/PMC186689/pdf/applmicro00014-0076.pdf

In it they use slants. I used LC. After my LC was heavily colonized I drew out 12mL into a syringe. The syringe was left to rest for an hour with the needle tip pointed downward. This allowed the suspended myc to fall to the bottom. After filling a BD vial with 9mL of sterile distilled water, I injected only the bottom 1mL of LC into the vial, taking care not to agitate the settled contents of the syringe. In this way the sugars of the LC were hopefully diluted enough and I had a dense delivery of myc into the vial.

I don't know if DI or distilled would make a difference. Sorry for mistyping which one I used in the earlier post. Hopefully someone else will chime in on this thread and comment on which water is preferable.


--------------------
“We need to make books cool again. If you go home with somebody and they don't have books, don't fuck them.”-John Waters

“Insanity is relative. It depends on who has who locked in what cage.”-Ray Bradbury

"The surest way to corrupt a youth is to instruct him to hold in higher esteem those who think alike than those who think differently."-Nietzsche

"There are no passengers on spaceship earth. We are all crew."-Marshall McLuhan

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Invisiblemicro
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Registered: 05/09/03
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Re: [Q] About isolating monoculture and its aging process [Re: ATXfungi]
    #21489713 - 04/01/15 08:42 PM (9 years, 1 month ago)

Quote:

ATXfungi said:
Measuring the amount of actives in your cubes is going to be outside your capabilities unless you have access to a university lab or private lab




Kovacs Reagent or Ehrlich's reagent

That said, I think he was referring more to viability and fruiting potential when he wrote that. He wasn't talking about P. cubensis specifically.

Personally, I haven't bothered. I got good enough yeilds from LC in bags of parboiled rice (I know it says to use brown but it's cheaper, fortified and doesn't stick together as much).

Also, use distilled water.

Edited by micro (04/01/15 08:50 PM)

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Offlinesstox
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Re: [Q] About isolating monoculture and its aging process [Re: micro]
    #21495907 - 04/03/15 10:26 AM (9 years, 1 month ago)

Thanks a lot for the answers and the linked paper. Very interesting to read. I'll try several techniques for comparing reasons.

Tbh I was writing about the potency. For viability and fruiting potiential I can isolate each culture and test the behaviour in growing, just like any other kind of mushroom. But I was wondering if there's an (easy) way to determine the amount of psilocybin/psilocin. But well, it could be worse than testing be eating them. ;-)

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