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InvisibleWall.E
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Re: Lurker finally posting question on “inconsistency of cultivation” [Re: Stipe-n Cap]
    #27046729 - 11/19/20 03:49 AM (3 years, 2 months ago)



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OfflineUzimyco
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Re: Lurker finally posting question on “inconsistency of cultivation” [Re: Wall.E]
    #27046851 - 11/19/20 07:56 AM (3 years, 2 months ago)

Thank you both. I will take your direction without question. That was always the intent.


For the sake of fun, I did want to use an illustration of what random info on the net provides, even so from this forum of course. (I am not referencing all the information you’ve all provided).

The attached image is the result of reading lots of tecs and doing things with precision.  You will see no such contamination as I rely heavily on sterilization with 257nm and 222nm radiation and do not waste much time with things such as bleach (I’m referencing inorganic items). Such radio sterilization virtually ensures the destruction of any contaminant. Even ones you cannot access or areas that cannot be seen.

This just 6 of 18 jars (remaining ones illustrate similar disparity) and this is my 4th unique run. Everything was done simultaneously. All variables were fixed. Things we’re introduced inside of a laminar flow hood. Spore was used from a communal vessel. Media was prepared all together and yet I followed instructions beyond the precision of the advice.  The photo illustrates what you already know. It however illustrates what was not discussed in stacks of literature.  You have all kindly filled in the blanks. I am only shocked at how significant time and effort in learning was stifled by incomplete information floating around.  Had I known before what I now know due to all of your patience, I would have avoided such mistakes.  This is the nature of learning. True it forced me to dig deeper and as a result I have moved to a greater level of understanding.  As long as I ultimately achieve the desired result, I’m ok with failure along the way. 

And if you’re wondering, yes I am well aware of the fact that I am absurdly verbose and blather incessantly. It is simply how I’m wired up when in a self education mode.  Such qualities have helped me in life equal to the damage they’ve done. I have accepted the benefits along with price one must pay.  Again although I’m sure you all do not require the reiteration: your patience and the information you’ve given is genuinely appreciated- it gives me comfort knowing a world dominated by selfish and unyielding people is offset by those who chose another path.  I’m actually quite typical in “real life”. Even “goofy” by the assessment of friends and contemporaries. But God did bestow an ability to write in the form you’re reading and allow me to articulate my thoughts in a professional manner.  You wouldn’t believe what contracts and legal documents look like when written by me...... they are quite ridiculous :wink:



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Thankfully the US Constitution ensures that my rights don't end where your feelings begin



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Offlinetravels_slightly
Tryptémon/Phenethylémon trainer
I'm a teapot User Gallery

Registered: 07/04/20
Posts: 47
Loc: The central California Co... Flag
Last seen: 3 years, 2 months
Re: Lurker finally posting question on “inconsistency of cultivation” [Re: Wall.E]
    #27046853 - 11/19/20 07:57 AM (3 years, 2 months ago)

I'm super grateful for threads like this. I also learned something, and I feel like p9hu7 just answered in a single post many of the things I was too shy to ask about out of fear I'd be punished for not UingTFSE my damn self, subsequently spending a whole lot of hours compiling conflicting information from which to draw my own (often incorrect) assumptions.

Thanks gang!


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General supporter of all things mystifying, specially when they happen to simultaneously be incriminating. ISO spores beyond my limited collection. I make and deal in many wares, perhaps we can exchange cool goodies :smile:


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InvisibleStipe-n CapMDiscord
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Re: Lurker finally posting question on “inconsistency of cultivation” [Re: Uzimyco] * 1
    #27046898 - 11/19/20 08:25 AM (3 years, 2 months ago)

Quote:

Uzimyco said:
I am a highly experienced horticulturalist in a wide array of complex genera




If you're already using a flowhood and have the above mentioned experience then I would wager that you have experience with plant tissue culture. Don't waste your time with spore solution and BRF. These have their place, of course,  they often serve as a point of contact for very new growers with limited exposure and experience.

Mush cult often seems very "sciency" and advanced when you're unfamiliar with the terminology.
Agar, petri dishes, sterilization/sterile technique, etc; they cause self doubt and hesitation. New growers often want to postpone their pursuit of such topics when they hear about what appears at first glance to be a shortcut to success with spores injected into a simple BRF recipe.

Cakes are often times not very noob friendly due to the complexity of relationships between competitive contamination found in spore syringes, the mushroom life cycle and it's relationship to it's environment. These subjects generate countless questions about their cakes being often contaminated, too dry/too wet, so on and so forth. These questions can only be answered with an experienced eye. Not very set and forget as a new grower may assume. It's not as easy as "hey, just going to squirt water into my jars and then become a mushroom Pablo Escobar in no time, brah"
New growers often find that once failure with that line of reasoning has forced them out of their comfort zone and into the warm welcoming arms of agar they realize there was nothing holding them back, it's actually super easy. Suddenly the world of mush opens up and success builds upon success. They always wish that they had started sooner.


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OfflineUzimyco
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Re: Lurker finally posting question on “inconsistency of cultivation” [Re: Stipe-n Cap]
    #27049990 - 11/21/20 07:42 AM (3 years, 2 months ago)

:wink: you are correct.

The fact that you’re familiar with TC already defines that your experience extends beyond mycology.

I freely admit that TC is something I had outstanding interest in at one time but that interest has waned. Obtaining quality aperical meristem tissue from rare almost one of a kind plants is a scary endeavor. Since it is “live material” I can’t “cheat” by using radiation sterilization.  It destroys all deoxyribonucleic acid 🧬 therefore rendering the tissue quite dead and no longer viable.  The extraction specifically of meristem tissue runs the heavy risk of killing a plant. When that plant is perhaps 1 of the 2 or 3 in the United States, it creates too much stress on myself and I’d virtually rather propagate my plants vegetatively.  Those kinds of mistakes are truly irreparable. Therefore I’ve chosen to leave TC to the true experts with vast resources of MS types of agar, specifically proven for a given genera. The bigger problem is that when dealing with really rare plants your opportunity to tweak media composition may well be the last mistake you make rendering the plant lost for the long term foreseeable future.  I’m sure you get the gist of what I mean.

In this case however such I’m in uncharted territory. For example - I can sterilize fern spore in a physan solution with quite good results. I have not tired this with fungi spore and I feel very sure it would be destructive. I have learned the hard way - fern spore is likely about as similar to muchroom spore as a car is to a dog. Spore is a term that structurally is dissimilar and fern spore maintains an outer layer that protects the delicate components. Fern spore is also exponentially larger than mushroom spore as fern spore can be individually viewed on paper whereas fungi spore are likely impossible to discern from a “pile”.

Also I use a wide array of endo and ecto mycorrhizae in horticulture. These are in fact fungi and I use them in tandem with products that introduce sub terranian pathinogenic support. I take it further by leveraging bacterium such as Streptomyces griseoviridis.  This bacterium however acts a fungicidal predator. It is known that it is effective as protection from fungi such as fusarium and it does promise to ignore non pathinogenic fungi. I’ve even considered trying such in mycology to see if it would attack cubensis spore or ignore it, while pedatorizing undesirable contaminants. There are other microorganisms I utilize but again I have relied far too much on horticulture knowledge when becoming interested in mycology and found them to be highly counterproductive. I’ve made cakes with the inclusion of things such as powdered black strap molasses as well as propolis since mycorrhizae flourish with it as a nutrient source. Furthermore using exotic combinations of much higher nutrition grains such as hemp flour yielded zero (and I mean zero results) so I backtracked to more traditional means.

Your posts and the posts of others in this thread have given me the education to stop conflating plants with fungi as the fact that they “grow” is roughly where the similarities end.  So yes. Again I’ve done a lot of messing with the hope that my “related knowledge” would supercharge my results. I have no shame in stating they have in fact, wildly done the opposite. I know when I’ve gone down the wrong road. Using control cultures and non control cultures illustrated that tried and traditional  media types categorically perform whereas exotic mixes with grain based superfoods don’t seem to be of any interest to mushrooms or at a minimum, the genera of mushrooms I’m working with.  As I said- even with great intent I suffered much failure and accepted the concept that I’m not looking to blaze new trail and my horticultural experience was quite worthless here.

Subsequent to the beginning of this thread and resulting from the information within, I have already acquired the necessary materials to make agar based dishes and that will be completed no later than tomorrow. Upon fruiting out the few cultures I have which appear to have hit full colonization, I will begin the agar process due to being able to collect fresh spore. I admit I feel empowered by the information I have been provided as failure is quite bothersome.

I thank you for the advice and I feel like I am on the correct path now. Time will tell but again, I feel a much greater understanding of many, many things that were not obvious to me.


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Thankfully the US Constitution ensures that my rights don't end where your feelings begin



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InvisibleStipe-n CapMDiscord
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Registered: 08/04/12
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Re: Lurker finally posting question on “inconsistency of cultivation” [Re: Uzimyco]
    #27050010 - 11/21/20 08:02 AM (3 years, 2 months ago)

Quote:

Uzimyco said:
. I admit I feel empowered by the information I have been provided as failure is quite bothersome.





Success is an error laden path. By learning what you shouldn't do you simultaneously learn what you should do. The fool who persists in their folly becomes wise. The only reason that I know not to do it that way is because I've tried it and failed in the exact same way.
Your failures are success because they make you wise.
Good luck.


Edited by Stipe-n Cap (11/21/20 08:11 AM)


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OfflineUzimyco
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Re: Lurker finally posting question on “inconsistency of cultivation” [Re: Stipe-n Cap]
    #27107091 - 12/25/20 08:46 AM (3 years, 1 month ago)

Well I am back again. Ill spare you some of the extraneous pitfalls I ran into, but nevertheless I finally had the ability to go spore straight to agar.

I did a couple of different strains on a couple of plates as a test but then decided to stop so that I could wait and see if I was doing it correctly. (This was recent. As in 2-4 days ago).

Admittedly my technique left much to be desired as the inoculation loops definitely did some cutting into the agar. I assumed that was ok but then again I feel like I can’t assume much of anything since my assumptions have proven wrong over and over.

To further make things exciting (I’m being sarcastic) and without a frame of reference I went a bit crazy with streaking the plates (lots of spore streaks) vs just the Z pattern. Again going back to the concept that more spore load - the better (yet again something I can not attest to being correct?).

So within the last couple of days something is certainly growing but I haven’t a clue if it’s right. It looks more like a translucent growth and not white mycelium at this point. Through radiation sterilizing it is virtually impossible to be just a containment as nothing I’ve ever done has exhibited “contamination type characteristics”.

So we come to the question at hand. Would anyone be able to provide any info as to timeframes of what should be determined by when?  I am incubating the plates at around 82 degrees in darkness 24 hrs a day.  I have read about 100 agar teks and they seem to detail much about prep and concept of spawning to bulk afterwards but not a lot on the process of agar colonization such as timeframes.  I do understand what I believe to be the aspect of selection a portion of healthy growth to replate as an isolate and grow on but that step would be after where I am at currently. Again just curious - what are you all incubating your plates at temp-wise and what is the timeframe that you generally make a determination as to what tissue you would want to isolate for growing on re-plated?

Many thanks to you all!


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Thankfully the US Constitution ensures that my rights don't end where your feelings begin



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