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OfflineLeafericson
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Who says you need agar and a Petri dish? King oyster.
    #24654767 - 09/24/17 12:20 AM (3 years, 7 months ago)


New to all of this so please pardon me if I'm out of place.
The above picture is of "king trumpet oyster"
It is slices of the stem in hard red winter wheat berries from 2 days ago.
From lots of research I have been led to believe that this form of cloning wasn't practical and close to impossible.
I said the hell with it and gave it a go.
As you can see it's growing quite rapidly.
When the mycelia has developed some more should I remove the stem cuttings to prevent rot?


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Offlinefalcon
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Re: Who says you need agar and a Petri dish? King oyster. [Re: Leafericson]
    #24655422 - 09/24/17 11:22 AM (3 years, 7 months ago)

No need to remove the cuttings, if it's going to rot, the contaminate that would make it do has most likely rode out on the mycelium.

It's not impossible at all, it's just for the most part impractical, it's easier to miss contaminates when you clone to grain.


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Re: Who says you need agar and a Petri dish? King oyster. (moved) [Re: Leafericson]
    #24655489 - 09/24/17 11:50 AM (3 years, 7 months ago)

This thread was moved from Advanced Mycology.

Reason:
Everyone can get lucky. Good luck doing that regularly


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InvisiblebodhisattaM
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Re: Who says you need agar and a Petri dish? King oyster. (moved) [Re: Leafericson]
    #24655494 - 09/24/17 11:51 AM (3 years, 7 months ago)

This thread was moved from Mushroom Cultivation.

Reason:
Oops meant to go here


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OfflineLeafericson
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Re: Who says you need agar and a Petri dish? King oyster. [Re: falcon]
    #24655581 - 09/24/17 12:17 PM (3 years, 7 months ago)

Quote:

falcon said:
It's not impossible at all, it's just for the most part impractical, it's easier to miss contaminates when you clone to grain.



Sorry I don't see the difference.
A sterilized petri dish of agar is sterile.
A sterilized jar of grain is sterile.
A slice of mushroom stem can be put in agar and thrive so why not in grain?


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OfflineLeafericson
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Re: Who says you need agar and a Petri dish? King oyster. (moved) [Re: bodhisatta]
    #24655590 - 09/24/17 12:19 PM (3 years, 7 months ago)

Quote:

bodhisatta said:
<img src='https://www.shroomery.org/forums/images/moved.gif'> This thread was moved from Advanced Mycology.

Reason:
Everyone can get lucky. Good luck doing that regularly



This was my first attempt at doing this so the odds of luck seem to be slim.
When I have the time I will replicate it to eliminate the chances of "luck".


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Re: Who says you need agar and a Petri dish? King oyster. (moved) [Re: Leafericson]
    #24655604 - 09/24/17 12:23 PM (3 years, 7 months ago)

Everyone has tried tissue culture clones right to grain before. We all still would do it as a regular thing if it actually worked well.

Not every clone will be clean so at least with agar your able to transfer away from the contamination then go to grain. Going straight to grain you just lose those jars


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OfflineLeafericson
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Re: Who says you need agar and a Petri dish? King oyster. (moved) [Re: bodhisatta]
    #24655618 - 09/24/17 12:27 PM (3 years, 7 months ago)

Quote:

bodhisatta said:
<img src='https://www.shroomery.org/forums/images/moved.gif'> This thread was moved from Mushroom Cultivation.

Reason:
Oops meant to go here



Considering not many people use this method and it actually worked I considered it to be in the realms of advanced mycology and hoped that it would be a beneficial alternative method to agar.
Not seeing many people doing this and I can't see why not.
If the medium is sterile and nutritious to the mycelium then why not use this method?
This would eliminate a step in the whole process of cloning.


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OfflineLeafericson
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Re: Who says you need agar and a Petri dish? King oyster. (moved) [Re: bodhisatta]
    #24655623 - 09/24/17 12:29 PM (3 years, 7 months ago)

Quote:

bodhisatta said:
Everyone has tried tissue culture clones right to grain before. We all still would do it as a regular thing if it actually worked well.

Not every clone will be clean so at least with agar your able to transfer away from the contamination then go to grain. Going straight to grain you just lose those jars



If the clone wasn't clean and healthy to start with I wouldn't attempt to use for cloning anyway. What would be the point? Wouldn't a contaminant on the stem transfer just the same to agar?


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OfflineMorePies
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Re: Who says you need agar and a Petri dish? King oyster. (moved) [Re: Leafericson]
    #24655667 - 09/24/17 12:44 PM (3 years, 7 months ago)

I have to culture away from contamination on at least 50 percent of my clones.


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Re: Who says you need agar and a Petri dish? King oyster. (moved) [Re: Leafericson]
    #24655683 - 09/24/17 12:50 PM (3 years, 7 months ago)

Quote:

Leafericson said:
Quote:

bodhisatta said:
Everyone has tried tissue culture clones right to grain before. We all still would do it as a regular thing if it actually worked well.

Not every clone will be clean so at least with agar your able to transfer away from the contamination then go to grain. Going straight to grain you just lose those jars



If the clone wasn't clean and healthy to start with I wouldn't attempt to use for cloning anyway. What would be the point? Wouldn't a contaminant on the stem transfer just the same to agar?



Clean and healthy sure but that doesn't mean you're so godly at agar work you'll never see a contamination. None of us would have any use for agar if you could simply get so good that you never got a contamination.

And yes it would transfer to the agar, thats the whole point. Then you could leave it behind and take only clean mycelium to a new dish to get a contamination free culture.

Its abundantly clear you're new to cloning and sterile work.


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Re: Who says you need agar and a Petri dish? King oyster. (moved) [Re: bodhisatta]
    #24655883 - 09/24/17 02:01 PM (3 years, 7 months ago)

Hey Leaferickson,

I think you're hearing the wrong message.

It is certainly possible to get desired mycelium to grow in unsterile conditions.

The problem is that you'll get sub-par yields ... possibly no yield at all depending on how aggressive the contaminants are.

What you're doing is great and is how knowledge advances ... by pushing the boundaries of our communal understanding we discover new things.  I applaud you!

This has, however, been tried in the past with limited success so is largely avoided now by working in sterile conditions and isolating strains on agar ... where it's much easier to identify contaminants.

Other things that do, in fact, work but are largely looked down upon are things like growing oyster mushrooms on freshly brewed coffee grounds ... you'll find that one all over the internet but it's almost never used by professional growers since it contaminates too easily and entire crops can be lost.

I started out this hobby with a similar approach and through hard-won knowledge via trial-and-error now agree that growing out on agar ultimately saves time and effort.  I hope that you'll benefit from my experience but understand if you need to find your own path.

Good luck and good growing,
adadada


Edited by adadada (09/24/17 02:02 PM)


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OfflineLeafericson
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Re: Who says you need agar and a Petri dish? King oyster. (moved) [Re: bodhisatta]
    #24656696 - 09/24/17 07:19 PM (3 years, 7 months ago)

Quote:

bodhisatta said:
Quote:

Leafericson said:
Quote:

bodhisatta said:

Its abundantly clear you're new to cloning and sterile work.



Your absolutely correct.
Strange thing though is that I have done many jars with grains and haven't had any contamination but the dam straw bags are very difficult for me to avoid contamination.
Seeing how many gourmet mushrooms are commercially grown from jars that maybe a better approach in my environment.


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OfflineLeafericson
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Re: Who says you need agar and a Petri dish? King oyster. (moved) [Re: adadada]
    #24656796 - 09/24/17 07:58 PM (3 years, 7 months ago)

Quote:

adadada said:
Hey Leaferickson,

I think you're hearing the wrong message.

It is certainly possible to get desired mycelium to grow in unsterile conditions.




I'm confused, the jar has been pressure cooked and as far as I can tell it is sterile since it was treated with the same respect as a petri dish would have been treated. The mushroom stem I wouldn't consider a high risk for contamination because it's the same method as if it was placed into agar.
Please explain so I can learn.

Quote:

adadada said:
The problem is that you'll get sub-par yields ... possibly no yield at all depending on how aggressive the contaminants are.



So far I've been fortunate and have had no contamination using grain in jars, however I can't seem to get very good results transferring to straw.

Quote:

adadada said:
What you're doing is great and is how knowledge advances ... by pushing the boundaries of our communal understanding we discover new things.  I applaud you!




Thanks, whether or not this method works out in the long run I appreciate the encouragement and if not as least I tried and may have learned something along the way.
Quote:

adadada said:
This has, however, been tried in the past with limited success so is largely avoided now by working in sterile conditions and isolating strains on agar ... where it's much easier to identify contaminants.




Yes I would imagine it would be much easier to see contamination on agar almost immediately. I would also imagine that contamination would spread much faster on agar and since there is less surface area the dish would be ruined. However with a jar of grain there is much more surface area for healthy mycelium to occupy, so I'm led to believe that if there was a spot of contamination the abundance of mycelium would have a better chance of taking over.
This maybe completely wrong but it makes sense in my head.
The bigest problem I could see is not seeing the contamination in the grain  like you would on agar and transferring that to your final substrate but wouldn't that be just the same if you started with agar and then went to grain?


Quote:

adadada said:
Other things that do, in fact, work but are largely looked down upon are things like growing oyster mushrooms on freshly brewed coffee grounds ... you'll find that one all over the internet but it's almost never used by professional growers since it contaminates too easily and entire crops can be lost.




Hahha so true. Coffee grows mold so fast.
However so companies are doing quite well using coffee.
Quote:

adadada said:
I started out this hobby with a similar approach and through hard-won knowledge via trial-and-error now agree that growing out on agar ultimately saves time and effort.  I hope that you'll benefit from my experience but understand if you need to find your own path.

Good luck and good growing,
adadada



I'm all ears, thanks for any and all advice positive or negative.
I have a tendency to follow the path less travelled finding my own way. I still highly respect the advice.


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Offlinefalcon
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Re: Who says you need agar and a Petri dish? King oyster. [Re: Leafericson]
    #24656973 - 09/24/17 09:10 PM (3 years, 7 months ago)

Yeah, sometimes if I'm cloning something wild I'll put it on grain as well as agar if it's something like oyster or Reishi and the weather is cool and the mushroom is young, if it takes I've a head start, if it doesn't I usually do this in a couple of 8 ounce jars and it's not that much of a hassle.


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OfflineLeafericson
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Re: Who says you need agar and a Petri dish? King oyster. [Re: falcon]
    #24657089 - 09/24/17 09:59 PM (3 years, 7 months ago)

Quote:

falcon said:
Yeah, sometimes if I'm cloning something wild I'll put it on grain as well as agar if it's something like oyster or Reishi and the weather is cool and the mushroom is young, if it takes I've a head start, if it doesn't I usually do this in a couple of 8 ounce jars and it's not that much of a hassle.



Wow so cool! So I assume you have success skipping the agar and going straight to grain  (?)
Pictures, links, threads, advice, any would be so very helpful.


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Re: Who says you need agar and a Petri dish? King oyster. [Re: Leafericson]
    #24658052 - 09/25/17 08:01 AM (3 years, 7 months ago)

O have had success, no pictures.


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OfflineFunguy182
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Re: Who says you need agar and a Petri dish? King oyster. (moved) [Re: Leafericson]
    #24661177 - 09/26/17 12:44 PM (3 years, 7 months ago)

I agree. With agar you can see the contamination and cut away and re transfer. I have done stem cutting inoculation and Everytime I have ended up with contamination. The mycelium did over take it but contaminated is contaminated.


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Re: Who says you need agar and a Petri dish? King oyster. (moved) [Re: Leafericson]
    #24665661 - 09/27/17 11:44 PM (3 years, 7 months ago)

Quote:

Leafericson said:
Yes I would imagine it would be much easier to see contamination on agar almost immediately. I would also imagine that contamination would spread much faster on agar and since there is less surface area the dish would be ruined. However with a jar of grain there is much more surface area for healthy mycelium to occupy, so I'm led to believe that if there was a spot of contamination the abundance of mycelium would have a better chance of taking over.
This maybe completely wrong but it makes sense in my head.





I think you are misunderstanding the purpose of agar work and how truly difficult it is to work sterile 100% of the time. One of the points of cloning on agar (as others have said) is so you can see contamination, transfer away from it and take only healthy culture and expand that onto grain or what have you. You don't let the mycelium on agar win out over the contamination, you remove healthy tissue from contamination until it's gone.

Quote:

Leafericson said:
The bigest problem I could see is not seeing the contamination in the grain  like you would on agar and transferring that to your final substrate but wouldn't that be just the same if you started with agar and then went to grain?




It's not the same thing because you are never transferring contaminated agar onto any sort of primary, secondary or final substrate. You would only ideally transfer healthy culture to these substrates from agar, which agar allows you to easily determine. Also, the problem with going directly from clone to grain is, if there is contamination you have no recourse and have to start over again, because it's near impossible to transfer healthy mycelium away from contamination on grain. It's not impossible to do straight-to-grain cloning, I don't think anyone has said that, but it's more likely to fail. I am actually doing this right now too, but it was because I didn't have access or time to make agar for what I wanted to clone.

I've worked with agar under ideal conditions, a 4' flowhood ran for 1 hour+, gloves, lab coat, mask, wiping everything down with ethanol, flame sterilizing utensils etc and still get a contamination rate of 10-20%. 100% sterile work is very difficult to achieve and using agar gives you a intermediary to work with.


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Re: Who says you need agar and a Petri dish? King oyster. (moved) [Re: DigitalRhizae]
    #24665714 - 09/28/17 12:12 AM (3 years, 7 months ago)

If contaminated you can always drop a grain on agar and try to clean. Just depends what the contaminate is. Logically agar is the first step as stated.


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