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InvisibleMycolorado
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Cleaning dirty cultures with tea agar * 9
    #23860431 - 11/23/16 12:12 PM (7 years, 2 months ago)

Just wanted to bring this to the attention of mush cult.  katbusa and Ferather have been experimenting with tea agar to clean up contaminated spawn and cultures.  I thought it would fit in over here as a great alternative to antibiotic agar...tea is readily available and not everyone wants to use antibiotics. Also, bacterial contams are often present when starting from prints and syringes, and can really be discouraging to those starting out.  I modified the original recipe to use less tea as the original amount seemed a bit excessive.  Here are a couple APE cultures and a malabar.  All had some degree of bacterial contam and are now growing clear.  Growth is very slow on this medium but will result in a clean culture. 
NOTE: spores will not germinate on this agar!



200ml H2O
4.25g black tea (2 bags)
2g ME
4g Agar



katbusa's tea agar thread
https://www.shroomery.org/forums/showflat.php/Number/23771470


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Re: Cleaning dirty cultures with tea agar [Re: Mycolorado]
    #23860444 - 11/23/16 12:17 PM (7 years, 2 months ago)

:takingnotes:


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Re: Cleaning dirty cultures with tea agar [Re: Mycolorado]
    #23860446 - 11/23/16 12:17 PM (7 years, 2 months ago)

:popcorn: interesting might have to give this a whirl.


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Re: Cleaning dirty cultures with tea agar [Re: concretelush] * 1
    #23860530 - 11/23/16 12:51 PM (7 years, 2 months ago)

Thats almost the exact amount of Tea I use in my T-Gel, 6g per 250g water.
Maximum stable Ph is 6, some lime balancing may be required.

Read more here and here. Thanks again Mycolorado.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather] * 2
    #23860533 - 11/23/16 12:52 PM (7 years, 2 months ago)

T-Gel is experimental, however does not need pressure cooking, only a microwave.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather] * 3
    #23860537 - 11/23/16 12:53 PM (7 years, 2 months ago)

Nice work. I know of some who also use activated charcoal to help inhibit bacteria. Just goes to show that antibiotics are the last resort not the first.


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Re: Cleaning dirty cultures with tea agar [Re: Pastywhyte] * 3
    #23860545 - 11/23/16 12:55 PM (7 years, 2 months ago)

The most potent is cinnamon, according to experts it's the most potent natural antibacterial there is.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #23860629 - 11/23/16 01:18 PM (7 years, 2 months ago)

This is something I've seen before n the only drawback is exactly what you warned about. .spores do not germinate on it and this goes for mold spores as well...these you simply drag to to the next plate or to your jars where they will germinate. Nice write up


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Re: Cleaning dirty cultures with tea agar [Re: cronicr]
    #23860645 - 11/23/16 01:22 PM (7 years, 2 months ago)



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Re: Cleaning dirty cultures with tea agar [Re: cronicr]
    #23860666 - 11/23/16 01:27 PM (7 years, 2 months ago)

Quote:

spores do not germinate on it and this goes for mold spores as well...these you simply drag to to the next plate or to your jars where they will germinate.




Do you think the mold spores are spread by the mycelium growth?
I never had this issue when I isolated healthy mycelium away from trichoderma.

If you transfer a piece of agar that is about 1,5 cm away from the "inoculation place" to another dish and make one more transfer for extra safety I doubt you have problems with trich in your jars.


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Re: Cleaning dirty cultures with tea agar [Re: enlightenment]
    #23860703 - 11/23/16 01:36 PM (7 years, 2 months ago)

That's just it though....without trich germinating how do you know if it's there


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Re: Cleaning dirty cultures with tea agar [Re: enlightenment] * 2
    #23860732 - 11/23/16 01:43 PM (7 years, 2 months ago)

The benefit for spores not germinating is the huge reduction of accidental contamination.
Both T-Gel and T-Agar can be assembled open air, not saying to do that.

I agree with enlightenment when it comes to Trich.

----

My first test sample of King oyster was infected with both bacillus cereus and trich.
The bacillus stalled after 24-48 hours, only mycelium grew, even the trich.

I isolated several sections using sterilized toothpicks.
Only the King oyster passed on to all the pegs.

   


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #23860739 - 11/23/16 01:46 PM (7 years, 2 months ago)

Here is the same sample a few days previously.

   


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #23860742 - 11/23/16 01:47 PM (7 years, 2 months ago)

I'm not saying there is anything wrong with it but just like anti biotic agar I would put through a standard recipe before going to grain spawn.


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Re: Cleaning dirty cultures with tea agar [Re: cronicr]
    #23860751 - 11/23/16 01:49 PM (7 years, 2 months ago)

That would be a problem if the tea dish is used to inoculate. Maybe I don't get you right or don't understand what you mean, sorry.

After further transfers it eliminates the risk IMO.

Example what I am thinking about:
- A tiny piece of an outdoor fruit (clone) to tea agar.
- Mycelium grows, trich or bacteria mostly stays in the middle of the plate
- Transfer a piece far away from the spot you placed the fruit sample to an agar dish without tea.
- Wait for healthy growth.
- Make another transfer
- Use this to inoculate grain

I give it a try with a contaminated Mexicana A. syringe I did not toss yet. I keep you posted. I hope it works because this syringe has cost time and nerves. :mad:

//edit

Quote:

cronicr said:
I'm not saying there is anything wrong with it but just like anti biotic agar I would put through a standard recipe before going to grain spawn.




That's what I tried to say :thumbup:


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Re: Cleaning dirty cultures with tea agar [Re: cronicr]
    #23860778 - 11/23/16 01:56 PM (7 years, 2 months ago)

Can bacteria piggy back on mycellium from plate to plate.  I really dont know what to do with these  P.Sandose plates

Transfer 2

Transfer 3

Do you think this could help?


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Re: Cleaning dirty cultures with tea agar [Re: enlightenment] * 1
    #23860786 - 11/23/16 01:58 PM (7 years, 2 months ago)

Sorry you guys are right, I have not mentioned the order of process I mentioned in my main post.

Dirty sample > T-Agar or T-Gel > Isolate > Transfer, agar > Validate > Spawn.


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Re: Cleaning dirty cultures with tea agar [Re: Edmunter]
    #23860792 - 11/23/16 02:02 PM (7 years, 2 months ago)

Quote:

Edmunter said:
Can bacteria piggy back on mycellium from plate to plate.  I really dont know what to do with these  P.Sandose plates

Transfer 2

Transfer 3

Do you think this could help?



Yes this would help or try sandwich it


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Offlineenlightenment
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Re: Cleaning dirty cultures with tea agar [Re: Edmunter]
    #23860794 - 11/23/16 02:02 PM (7 years, 2 months ago)

Quote:

Edmunter said:
Can bacteria piggy back on mycellium from plate to plate.  I really dont know what to do with these  P.Sandose plates

Transfer 2

Transfer 3

Do you think this could help?




You should give it a try. It is hard to get rid off bacteria. I gave up some dishes because of massive problems. Made up to 5 transfers. Bacteria is harder to defeat than trich because bacteria can grow very fast. It is hard to isolate healthy mycelium away. Antibiotic agar sounds like a godsend for this purpose.

Try it and enrich this thread with your experience.


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Re: Cleaning dirty cultures with tea agar [Re: cronicr]
    #23860798 - 11/23/16 02:04 PM (7 years, 2 months ago)

I will try this tek and post here, maybe along side a sandwich to compare


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Re: Cleaning dirty cultures with tea agar [Re: Edmunter]
    #23860882 - 11/23/16 02:38 PM (7 years, 2 months ago)

Sandwich works perfectly, katbusa has thankfully tested that method, check the results here.
This was however the older first attempt recipe katbusa developed, credit due.

Both myself and Mycolorado have updated the ratio.


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Re: Cleaning dirty cultures with tea agar [Re: cronicr] * 1
    #23860903 - 11/23/16 02:46 PM (7 years, 2 months ago)

Quote:

cronicr said:
I'm not saying there is anything wrong with it but just like anti biotic agar I would put through a standard recipe before going to grain spawn.



Sorry, yep, after it's looking clean on the tea agar, it should be transferred to and grown out on your standard recipe.


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Re: Cleaning dirty cultures with tea agar [Re: Mycolorado]
    #23860908 - 11/23/16 02:48 PM (7 years, 2 months ago)

You mentioned very slow growth.
I'm assuming there's a reason for the malt extract but for arguments sake, say you took out the 2g ME from the mix. Would the tea have enough nutes to support mycelium?


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Re: Cleaning dirty cultures with tea agar [Re: ComebackKid] * 1
    #23860931 - 11/23/16 02:57 PM (7 years, 2 months ago)

My guess is yes.  Pretty sure Ferather's recipe doesn't use the malt extract.  This is definitely a work in progress and I would encourage experimentation with different percentages, though Ferather is doing a pretty good job dialing it in.


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Re: Cleaning dirty cultures with tea agar [Re: ComebackKid]
    #23860933 - 11/23/16 02:57 PM (7 years, 2 months ago)

Quote:

ComebackKid said:
Would the tea have enough nutes to support mycelium?




Yes, dry tea raw per 100g is richer than 100g dry rye (change to 100g).
The slower growth is due to Ph, not nutrition or energy.



Note the N and protein.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #23860992 - 11/23/16 03:15 PM (7 years, 2 months ago)

Potential energy and  N sources (incomplete) for tea.

Polyphenols, varied: [ C6 | H5 | O1 ] x.
Cellulose, varied: [ C6 | H10 | O5 ] x.

Theophylline: C7 | H8 | N4 | O2.
Theobromine: C7 | H8 | N4 | O2.
Caffeine: C8 | H10 | N4 | O2.
Theaflavin: C29 | H24 | O12.
Tannins: C76 | H52 | O46.
Catechin: C15 | H14 | O6.
Carotene: C40 | H56.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #23861008 - 11/23/16 03:21 PM (7 years, 2 months ago)

Quote:

Ferather said:
Quote:

ComebackKid said:
Would the tea have enough nutes to support mycelium?




Yes, dry tea raw per 100g is richer than 100g dry rye (change to 100g).
The slower growth is due to Ph, not nutrition or energy.



Note the N and protein.



Wouldn't a richer sub colonize slower


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Re: Cleaning dirty cultures with tea agar [Re: cronicr]
    #23861021 - 11/23/16 03:24 PM (7 years, 2 months ago)

Funny you say that, that also needs further testing, however various elements and vitamins improve growth speed.


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Re: Cleaning dirty cultures with tea agar [Re: cronicr]
    #23861024 - 11/23/16 03:24 PM (7 years, 2 months ago)

Quote:

cronicr said:
Quote:

Ferather said:
Quote:

ComebackKid said:
Would the tea have enough nutes to support mycelium?




Yes, dry tea raw per 100g is richer than 100g dry rye (change to 100g).
The slower growth is due to Ph, not nutrition or energy.



Note the N and protein.



Wouldn't a richer sub colonize slower




That is my thoughts as well.


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Re: Cleaning dirty cultures with tea agar [Re: Pastywhyte]
    #23861047 - 11/23/16 03:31 PM (7 years, 2 months ago)

Indeed it needs testing, something I will try. Yeast nutrients.

   

Watch what happens, given the right ratio.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #23862618 - 11/24/16 05:45 AM (7 years, 2 months ago)

Hi Does anyone have a link newer than this or is this fine https://www.shroomery.org/forums/showflat.php/Number/2894662


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Re: Cleaning dirty cultures with tea agar [Re: Mycolorado]
    #23862620 - 11/24/16 05:47 AM (7 years, 2 months ago)

Quote:

Mycolorado said:
Just wanted to bring this to the attention of mush cult.  katbusa and Ferather have been experimenting with tea agar to clean up contaminated spawn and cultures.  I thought it would fit in over here as a great alternative to antibiotic agar...tea is readily available and not everyone wants to use antibiotics. Also, bacterial contams are often present when starting from prints and syringes, and can really be discouraging to those starting out.  I modified the original recipe to use less tea as the original amount seemed a bit excessive.  Here are a couple APE cultures and a malabar.  All had some degree of bacterial contam and are now growing clear.  Growth is very slow on this medium but will result in a clean culture. 
NOTE: spores will not germinate on this agar!



200ml H2O
4.25g black tea (2 bags)
2g ME
4g Agar



katbusa's tea agar thread
https://www.shroomery.org/forums/showflat.php/Number/23771470




Does ordinary house hold PG tips work with this?  Uk tea.


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Re: Cleaning dirty cultures with tea agar [Re: Edmunter]
    #23862711 - 11/24/16 06:52 AM (7 years, 2 months ago)

Quote:

Edmunter said:
Hi Does anyone have a link newer than this or is this fine https://www.shroomery.org/forums/showflat.php/Number/2894662




That link is fine but you can skip the peroxide and you don't need gentamiacin to make it work.


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Re: Cleaning dirty cultures with tea agar [Re: Pastywhyte]
    #23862720 - 11/24/16 06:56 AM (7 years, 2 months ago)

Quote:

Pastywhyte said:
Quote:

Edmunter said:
Hi Does anyone have a link newer than this or is this fine https://www.shroomery.org/forums/showflat.php/Number/2894662




That link is fine but you can skip the peroxide and you don't need gentamiacin to make it work.




Wait, so im just making an agar plate with no added nutes?


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Re: Cleaning dirty cultures with tea agar [Re: Edmunter]
    #23862724 - 11/24/16 07:01 AM (7 years, 2 months ago)

You need nutes. Just not peroxide or gentamiacin.


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Re: Cleaning dirty cultures with tea agar [Re: Pastywhyte]
    #23862752 - 11/24/16 07:14 AM (7 years, 2 months ago)

So just a normal PDA recipe is fine.  Its not clear in that write up.


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Re: Cleaning dirty cultures with tea agar [Re: Edmunter]
    #23862754 - 11/24/16 07:14 AM (7 years, 2 months ago)

Hey Ed, any tea should work (I would stick to unflavored).  Green tea should also be investigated...I think katbusa was testing it.  Hopefully, tea agar will prove effective in helping clean up a variety of contams and peroxide and antibiotics can be avoided.


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Edited by Mycolorado (11/24/16 07:34 AM)


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Re: Cleaning dirty cultures with tea agar [Re: Mycolorado]
    #23862791 - 11/24/16 07:30 AM (7 years, 2 months ago)

I am a very modern man and only have fruit teas like passion fruit and mango....... Joking... Thanks Myc I may try both. 

Do you think my P.Sandose looks like it has bacteria as I havent been able to shift the weird growth.  Looks like you turned up just in time.


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Re: Cleaning dirty cultures with tea agar [Re: Edmunter]
    #23862805 - 11/24/16 07:38 AM (7 years, 2 months ago)

I use the extra cheap supermarket stuff. It's more like flour than leaves.
Any unmodified (flavored, e.g. added lemon) raw tea leaves.

For extra benefits balance to Ph 6 using lime.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #23862811 - 11/24/16 07:41 AM (7 years, 2 months ago)

Quote:

Ferather said:
I use the extra cheap supermarket stuff. It's more like flour than leaves.
Any unmodified (flavored, e.g. added lemon) raw tea leaves.

For extra benefits balance to Ph 6 using lime.





Now youve gone and done it,  The missus wants to know WTF I want litmus paper for........ I blamed you......


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Re: Cleaning dirty cultures with tea agar [Re: Edmunter]
    #23863053 - 11/24/16 09:30 AM (7 years, 2 months ago)

Fuck yeah I might go get some tea now and keep it in my cabinet just to try this out when the time comes


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Re: Cleaning dirty cultures with tea agar [Re: Edmunter]
    #23863061 - 11/24/16 09:34 AM (7 years, 2 months ago)

Quote:

Edmunter said:
Now youve gone and done it,  The missus wants to know WTF I want litmus paper for........ I blamed you......



Lol, sorry. They are dirt cheap, got 160 for £1.50 free P&P. Cut them in half, you get 320.


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Re: Cleaning dirty cultures with tea agar [Re: Leftfield420]
    #23863176 - 11/24/16 10:18 AM (7 years, 2 months ago)

Quote:

Leftfield420 said:
Fuck yeah I might go get some tea now and keep it in my cabinet just to try this out when the time comes




Are you a heathen,  U cant live in England unless you have tea in your cupboard


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Re: Cleaning dirty cultures with tea agar [Re: Edmunter]
    #23863437 - 11/24/16 11:52 AM (7 years, 2 months ago)

I like this!

Should of prepped my recent plates with some cinnamon/tea, oh well. :facepalm:


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Re: Cleaning dirty cultures with tea agar [Re: Teemo 6T3]
    #23863935 - 11/24/16 03:36 PM (7 years, 2 months ago)

I have a request Mycolorado, if you have the time. Could you try a starch free version?
You can transfer from one of those T-Agars you have now, Ph 6 via lime.

You shouldn't need to pressure cook. Thanks again.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #23864081 - 11/24/16 04:40 PM (7 years, 2 months ago)

Of course. Out of town on holiday but will put some together this weekend.


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Re: Cleaning dirty cultures with tea agar [Re: Mycolorado]
    #23864091 - 11/24/16 04:44 PM (7 years, 2 months ago)

Top man, no rush whatsoever  :thumbup:


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #23864222 - 11/24/16 06:03 PM (7 years, 2 months ago)

:takingnotes:


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Re: Cleaning dirty cultures with tea agar [Re: Edmunter]
    #23864244 - 11/24/16 06:11 PM (7 years, 2 months ago)

Man I don't drink tea...and don't live in england..


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Re: Cleaning dirty cultures with tea agar [Re: Leftfield420]
    #23864930 - 11/24/16 11:16 PM (7 years, 2 months ago)

:takingnotes:


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Re: Cleaning dirty cultures with tea agar [Re: Leftfield420]
    #23865400 - 11/25/16 07:49 AM (7 years, 2 months ago)

Quote:

Leftfield420 said:
Man I don't drink tea...and don't live in england..





Then u know how lucky u are.......


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Re: Cleaning dirty cultures with tea agar [Re: Edmunter]
    #23880725 - 11/30/16 05:48 AM (7 years, 1 month ago)

Ok so I took this



and made 2 sandwiches




Next minute im free(next week) I will make a Tea plate to compare.  Im still not sure this plate is contaminated but its worth a try.


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Re: Cleaning dirty cultures with tea agar [Re: Edmunter]
    #23905951 - 12/08/16 07:29 AM (7 years, 1 month ago)

should i just boil the bags or throw the tea powder on the mix ?


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Re: Cleaning dirty cultures with tea agar [Re: Neomorph]
    #23906013 - 12/08/16 08:02 AM (7 years, 1 month ago)

I use the bag method, you can do leaves and strain. You want the solubles not the leaves.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #23906062 - 12/08/16 08:26 AM (7 years, 1 month ago)

just boiled the bags with water downt to 200 ml :smile:


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Re: Cleaning dirty cultures with tea agar [Re: Neomorph]
    #23906215 - 12/08/16 09:59 AM (7 years, 1 month ago)

Here's a malabar culture after cleaning with tea agar.  It was pretty dirty from the syringe and was growing very poorly on each transfer from the original plate.  This is the first transfer from tea and looks great.


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Re: Cleaning dirty cultures with tea agar [Re: Mycolorado]
    #23906244 - 12/08/16 10:12 AM (7 years, 1 month ago)

Mold will grow if spores get transfered onto tea agar (waited to long to transfer from a plate with mold), but it does keep bacteria away for the most part...I did a controlled experiment using 1tea agar plate, and 1 PDA plate: took mouth swabs and streaked each plate on 11/27 I have visible bacterial growth in the PDA plate, and not anything that I can see with my naked eye on the tea agar plate


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Re: Cleaning dirty cultures with tea agar [Re: Leftfield420]
    #23906476 - 12/08/16 11:21 AM (7 years, 1 month ago)

Quote:

Leftfield420 said:
Mold will grow if spores live mold gets transfered onto tea agar.




:smile:


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #23906486 - 12/08/16 11:24 AM (7 years, 1 month ago)

Mold spores will still germinate in your spawn tho:shrug:


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Re: Cleaning dirty cultures with tea agar [Re: ComebackKid]
    #23906502 - 12/08/16 11:28 AM (7 years, 1 month ago)

Working on that one, but normally yes. But not from cooking, mis-handling, filters and so on.


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Re: Cleaning dirty cultures with tea agar [Re: ComebackKid]
    #23906507 - 12/08/16 11:28 AM (7 years, 1 month ago)

I think we discussed above you need to make one more transfer to clean standard media


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #23906512 - 12/08/16 11:29 AM (7 years, 1 month ago)

How's that grain spawn test going btw?


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Re: Cleaning dirty cultures with tea agar [Re: ComebackKid]
    #23906525 - 12/08/16 11:32 AM (7 years, 1 month ago)

This is not my thread, but so far positive results. Katbusa is beating me to it though.


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Re: Cleaning dirty cultures with tea agar [Re: ComebackKid]
    #23906538 - 12/08/16 11:37 AM (7 years, 1 month ago)

Good to know... The mold must have piggybacked my mycelium then? Because I transfered from the furthest point from the visible mold...


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #23906548 - 12/08/16 11:39 AM (7 years, 1 month ago)

Im rather fond of using the property of the agar as another member stated

Wish I coukd remember the link ill dig it up..

I noticed when using old dried out (1-2month old) plates for cloning my bacterial contamination rates plummeted. I never looked back. Just keep petris in zip lock bags.

Another member started a thread on how he likes to increase the amount of agar added to the water. Which in effect makes a much drier agar surface. Same effect I was gaining just a diff way of going about it.

I haven't experimented with this type of method and cleaning up cultures but cloning it does wonders.

I like this idea tho,  maybe start your spores on regular agar then throw mycelium onto tea agar for ur antibiotic agar effect. Then back to regular agar before u go to grains


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Re: Cleaning dirty cultures with tea agar [Re: Raven44]
    #23906578 - 12/08/16 11:47 AM (7 years, 1 month ago)

Quote:

Leftfield420 said:
Good to know... The mold must have piggybacked my mycelium then? Because I transfered from the furthest point from the visible mold...



I have done 4 spawn samples that have no visible mold, and then mold coming from it.


Quote:

Raven44 said:
........
I like this idea tho,  maybe start your spores on regular agar then throw mycelium onto tea agar for ur antibiotic agar effect. Then back to regular agar before u go to grains



Thank you for that information, you can use this agar however you like.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24078721 - 02/09/17 11:36 PM (6 years, 11 months ago)

wondering if anybodies been continuing to test this idea? noticed ferathers thread on the subject is no longer with us so thought I'd ask here


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Re: Cleaning dirty cultures with tea agar [Re: Psilosopherr]
    #24078933 - 02/10/17 05:21 AM (6 years, 11 months ago)

I have a jar of tea agar I've had for months with no apparent bacteria or mold in it....and I still have the tea agar petri I mouth swabbed... Little to no growth on it...definently curves the growth of bacteria.. If mold spores are introduced they will grow on tea agar though


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Re: Cleaning dirty cultures with tea agar [Re: Leftfield420]
    #24420644 - 06/20/17 01:58 PM (6 years, 7 months ago)

this is really cool mycolorado. are you still using the tea agar? i think im going to try this recipe out. also, the color really pops the mycelia. bookmarked and im adding this to my reference list.


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Re: Cleaning dirty cultures with tea agar [Re: eatyualive]
    #24420731 - 06/20/17 02:32 PM (6 years, 7 months ago)

Hey eat!  I haven't used it since my initial dirty cultures from syringe.  Both my Malabar and APE were REALLY dirty.  I think it would be a great tool for cleaning up tissue cultures from wild specimen as well and will be giving it a try this summer with some wild oyster clones.


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Re: Cleaning dirty cultures with tea agar [Re: Mycolorado]
    #24422111 - 06/20/17 09:15 PM (6 years, 7 months ago)

I had success cleaning a dirty culture with this method, I used green tea.

Won't need it very often in my career but I'd definitely use it again


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Re: Cleaning dirty cultures with tea agar [Re: Psilosopherr]
    #24422121 - 06/20/17 09:19 PM (6 years, 7 months ago)

Green tea worked? I might give that a try myself. Not pinning my hopes or nothing but I got a culture that I may give this a go with anyway.


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Re: Cleaning dirty cultures with tea agar [Re: Pastywhyte] * 1
    #24423238 - 06/21/17 08:58 AM (6 years, 7 months ago)

It works in the way of alternative carbon sources (non-sugar), 100g of black tea has 3g natural sugar no starch.
As it stands without the trace carbohydrates, it's very nutritional and soluble, even cheap black tea.

The natural carbon containing acids breakdown at about ph 6.5, so pH 7-7.5 is ideal.
You can still get some bacteria but total activity should be reduced overall.

Here is some composition links (external), Tea, Tea to drink.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather] * 1
    #24461944 - 07/06/17 12:44 PM (6 years, 6 months ago)

I setup another recipe, sugar and starch free. I am using cheap black tea, about 80p for 250g.
My setup is using a cellulose bacteria (also eats starch and sugar) as the infection.

The recipe and further explanation can be found in katbusa's thread here.
As a side note cubensis will transfer to a wood toothpick peg.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather] * 1
    #24466933 - 07/08/17 06:43 AM (6 years, 6 months ago)

Here is an export of another thread I made, related to a carbon discussion.
You can use the produced enzymes to create a controlled media.


Digestive enzymes:

Amylase - starch  []  Laccase - lignin  []  Cellulase - cellulose  []  Xylanase - hemicellulose  []  Protease - proteins.

Mycelium should only cleave the bonds of carbohydrate's and hydrocarbon's, C-H bonded units.
It should not for example degrade materials of a C-C bonding, such as coal or diamonds.

However there may be specialized mycelium that may, it will be rare.

----

Carbon-Hydrogen targeting:

Amylase, Laccase, Cellulase, Xylanase


Carbon + Nitrogen targeting:

Protease

----

Example carbon materials:

Starch, Sugar, Cellulose, Hemicellulose, Lignin, Tannins, Bicarbonates.


Example nitrogen materials:

Proteins, Vitamins, Caffeine, Ureic, Ammonical.

----

Side note:

Laccase will decay many carbon materials.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24467057 - 07/08/17 08:02 AM (6 years, 6 months ago)

It would be very cool if green tea works. id love to see how a green tea plate would look.

damnit you guys are making me do work now. i have to try this out.


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Re: Cleaning dirty cultures with tea agar [Re: eatyualive] * 1
    #24467077 - 07/08/17 08:14 AM (6 years, 6 months ago)

I believe katbusa tested green tea with positive results, although a small amount of malt extract was used.
He did note the same oxidization effect around the mycelium, though less coloured, more brown.

Technically speaking green tea extract should be richer, and closer to natural.
The mycelium should have no issues oxidizing materials with laccase.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather] * 1
    #24467107 - 07/08/17 08:28 AM (6 years, 6 months ago)

Identification of contamination (where CaCO3 is being used) is fairly easy, bacteria: yellow, yeast: creamy.
Mycelium, this includes molds (they are a mycelium too!), look normal when growing.

   


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Re: Cleaning dirty cultures with tea agar [Re: Ferather] * 1
    #24467111 - 07/08/17 08:31 AM (6 years, 6 months ago)

I suggest going sugar and starch free, or at least half normal ME dose.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather] * 2
    #24467226 - 07/08/17 09:22 AM (6 years, 6 months ago)

I also suggest using a sterilized wood toothpick to isolate-transfer mycelium, as it helps restrict contamination.
You can see my signature (below) to see my toothpick and cellulose agar guides, as a reference.

The above culture, overpowered present contamination eventually, a sample was sent.

Here are three posts of the results of the sent sample: One, Two, Three.
Also, here is another case of a peg transfer from UK > US.

   


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Re: Cleaning dirty cultures with tea agar [Re: Ferather] * 1
    #24467229 - 07/08/17 09:24 AM (6 years, 6 months ago)

   


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24467300 - 07/08/17 09:51 AM (6 years, 6 months ago)

I was just looking at this the other day Ferather. Love it! :thumbup:


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Re: Cleaning dirty cultures with tea agar [Re: eatyualive]
    #24467388 - 07/08/17 10:24 AM (6 years, 6 months ago)

All you need is metal tweezers, cheap and easy to get, easy to clean or sterilize.
Transfers take seconds, the area of contact is away from the agar.

You can also replace removed pegs with new pegs.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24468063 - 07/08/17 04:30 PM (6 years, 6 months ago)

Ok made up some no pour plates today.

used

1 cup brewed tea from the tea maker.(6 grams tea-3 bags,  4 cups water)
1 tablespoon or close to 6 grams MEA.

after the tea brew, cooked it on medium heat and added the agar. stirred until the mix was consistent. then poured into plates.

pced for 30 minutes at 15-17 psi.







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Re: Cleaning dirty cultures with tea agar [Re: eatyualive]
    #24469864 - 07/09/17 11:27 AM (6 years, 6 months ago)

Do you know how much tea that was? Sorry I work by weighing, an estimate is fine, thnx.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24470342 - 07/09/17 02:38 PM (6 years, 6 months ago)

Did I mention that I used just hot water (no pressure cook) and I assembled everything open air.
If I do the same with an agar recipe with sugar or starch I need to PC, and get mold.

I still have no contamination, and I'm adding transfer pegs tomorrow.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24470998 - 07/09/17 07:11 PM (6 years, 6 months ago)

4 cups water was brewed with 6 grams tea.1 cup of that water was used for the plates.


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Re: Cleaning dirty cultures with tea agar [Re: eatyualive]
    #24471831 - 07/10/17 06:17 AM (6 years, 6 months ago)

Ok, thank you.

:takingnotes:


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24471846 - 07/10/17 06:32 AM (6 years, 6 months ago)

Great improv with the tooth picks, I think use this method also with pieces of straw, or stinging nettle, or whatever substrate you want to train the myc for. I suddenly think of myclial expansion as a pre-school for the myc and every bit of substrate is a teaching opportunity. I like to put bits of whatever substrates or spawn mediums in the agar that not only select out the strains but to give them a genetic memory of how to deal with these subs in the future. I theorize that you can train mycelium to deal with all sorts of conditions. By retaining a preserved culture of the original isolate I suspect you may be able to inoculate the fresh mycelium with the mycelial memory of the same strain after it has been through a lot, Thereby maintaining vigor and the learned responses to different situations. Think of it as a Grandad myc passing it's Knowledge to this grand son who has a long brilliant futre ahead of it and all of the infinite potential in the universe.


Edited by Lennybernadino (07/10/17 06:34 AM)


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Re: Cleaning dirty cultures with tea agar [Re: Mycolorado]
    #24471898 - 07/10/17 06:59 AM (6 years, 6 months ago)

:thumbup:


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Re: Cleaning dirty cultures with tea agar [Re: Morelman]
    #24471911 - 07/10/17 07:05 AM (6 years, 6 months ago)

This is awesome. Thanks for sharing. This should make clean up easier.

@ ferather u seriously do not pc with this method?


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Re: Cleaning dirty cultures with tea agar [Re: Morelman]
    #24471913 - 07/10/17 07:06 AM (6 years, 6 months ago)

Thanks everyone, this thread is open so feel free to experiment, share and discuss the topic.

@EdiblesOnly, the recipe in my signature, nope no pressure cook, open air.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24471949 - 07/10/17 07:22 AM (6 years, 6 months ago)

:takingnotes:


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Re: Cleaning dirty cultures with tea agar [Re: Mycolorado]
    #24472012 - 07/10/17 07:59 AM (6 years, 6 months ago)

Neat, thank you for bringing this to my attention. Might have to give it a whirl!


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Re: Cleaning dirty cultures with tea agar [Re: MagikShrooms]
    #24472051 - 07/10/17 08:26 AM (6 years, 6 months ago)

No problem. The carbon-nitrogen ratio of the T-Gel agar is about 30:1, roughly, using non sugar carbon.

List of carbon sources, remaining required nutrients from the tea and gelatin.
The nutrients come in a natural and organic water soluble format.


Calcium bicarbonate: [ Ca | C | H | O3 ] 2.
Proteins, varied: [ Cx | Hx | Ox | Nx ] x.
Polyphenols, varied: [ C6 | H5 | O ] x.

Theophylline: C7 | H8 | N4 | O2.
Theobromine: C7 | H8 | N4 | O2.
Caffeine: C8 | H10 | N4 | O2.
Theaflavin: C29 | H24 | O12.
Tannins: C76 | H52 | O46.
Catechin: C15 | H14 | O6.
Carotene: C40 | H56.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24472372 - 07/10/17 11:31 AM (6 years, 6 months ago)

I kinda worked the recipe into the way I usually do my no pour plates. Since I had mea I just assumed 6 grams total.

Transfers are looking good today. I'll get pics up in a few days. I also wanted to see if a slight variation would work. I like mixing it up. That amount I stated above poured 12 glad rounds.

The soul intent of me using this was for an old cubie called matias Romero. First swipe looks clean but wanted to try it out. I'd also like to spread the spores on this matias Romero. Its one of my old favorites from 99-2000.


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Re: Cleaning dirty cultures with tea agar [Re: eatyualive] * 1
    #24472498 - 07/10/17 12:31 PM (6 years, 6 months ago)

Makes sense, nutritional variation should allow it to detect-digest more things, or if you like, use more genetic blueprints.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather] * 1
    #24472644 - 07/10/17 01:31 PM (6 years, 6 months ago)

Here are today's images of my T-Gel agar, no pressure cook, open air assembly, no contamination.

           


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24473138 - 07/10/17 04:15 PM (6 years, 6 months ago)

There's contamination it's just not germinating/growing. It's not like you're keeping it out, you're keeping it at bay

Who's moved off of open air tea agar back to sterilised agar?
Or taken open air agar and inoculated sterilized spawn?

Is there a way to accomplish that or do you take you open air agar to open air-able grows?


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Re: Cleaning dirty cultures with tea agar [Re: bodhisatta] * 3
    #24474960 - 07/11/17 10:58 AM (6 years, 6 months ago)

Both agar and grain transfer without infection, from open air transfers, here is the post I already provided.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24475057 - 07/11/17 11:35 AM (6 years, 6 months ago)

Yea I've seen that already wondering if you actually grew some mushrooms though like to prove that grain transfer worked well


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Re: Cleaning dirty cultures with tea agar [Re: bodhisatta]
    #24475079 - 07/11/17 11:42 AM (6 years, 6 months ago)

You mean like Mattisfat's king oyster log? As far as TA you will need to ask him. Here is WBS, then WBS to WL-Tek.

                 

                 


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24475095 - 07/11/17 11:47 AM (6 years, 6 months ago)

Oysters on WL tek isn't compelling they already grow on cellulose product


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Re: Cleaning dirty cultures with tea agar [Re: bodhisatta]
    #24475123 - 07/11/17 11:55 AM (6 years, 6 months ago)

What do you think coco coir is composed of?


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24475126 - 07/11/17 11:57 AM (6 years, 6 months ago)

Are those the centrifugal plastic vials you mentioned in your initial post?


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Re: Cleaning dirty cultures with tea agar [Re: MagikShrooms]
    #24475137 - 07/11/17 12:00 PM (6 years, 6 months ago)

Yes, they are 2ml centrifugal vials, which is a perfect size for 1/2 a toothpick.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24475248 - 07/11/17 12:41 PM (6 years, 6 months ago)

Coir isnt the food the spawn is :shrug: I don't think I've seen a straight coir grow...


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24475262 - 07/11/17 12:45 PM (6 years, 6 months ago)

Awesome, thank you. They look like they can be very handy. Like single use inoculation media,


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Re: Cleaning dirty cultures with tea agar [Re: MagikShrooms]
    #24475687 - 07/11/17 03:32 PM (6 years, 6 months ago)

I missed the open air knock bit as well

So you boil your grains in tea or..? Bulk sub too?


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Re: Cleaning dirty cultures with tea agar [Re: Psilosopherr]
    #24475727 - 07/11/17 03:48 PM (6 years, 6 months ago)

Quote:

bodhisatta said:
Coir isnt the food the spawn is :shrug: I don't think I've seen a straight coir grow...




No, that's not really what is happening, the spawn acts as the nutrient source in that scenario, in the same way as adding grain flour to verm.
Surely this has been tried, so what is better, 50g grain spawn + 50g dry verm + water, or, 50g grain spawn + 50g dry coir + water.

If the verm was better, surely you would prefer this over coir, as the verm will not contaminate, unless you add things.
Coco coir is considered "cold" due to lack of nutrients, composed mostly of cellulose, hemicellulose and lignin.


Quote:

Psilosopherr said:
I missed the open air knock bit as well

So you boil your grains in tea or..? Bulk sub too?




??


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24475732 - 07/11/17 03:50 PM (6 years, 6 months ago)

If verm was cheaper than coir I probably would use all verm substrate..


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Re: Cleaning dirty cultures with tea agar [Re: bodhisatta]
    #24475769 - 07/11/17 04:02 PM (6 years, 6 months ago)

So a side by side has been done, considering you could measure a small amount, say 100g.
Can I have the information for this side-by-side, and relational recipe's.

Thanks.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24476195 - 07/11/17 07:02 PM (6 years, 6 months ago)

I have done rez effect a few times. It was nearly identical to the coir. If verm was cheaper than coir I would use it in far larger amounts. The only thing is it does best cased with something other than verm. But as far as yeild when you do case with a mix, it's pretty well the same.


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Re: Cleaning dirty cultures with tea agar [Re: Pastywhyte]
    #24490023 - 07/17/17 06:23 PM (6 years, 6 months ago)

Finally giving this a shot

Just mixed 2 grams of red rose orange pekoe into munches 1 2 3 pda recipe.
Can't wait to run my cultures through it. :thumbup:

Lmao my dumbass dropped a couple drops of green food colouring in there so I could differentiate between the tea agar and my regular blue agar plates.
Completely forgot that tea turns fucking brown anyway :lol::facepalm3:


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Re: Cleaning dirty cultures with tea agar [Re: ComebackKid]
    #24490172 - 07/17/17 07:39 PM (6 years, 6 months ago)

:thumbup:.

I had to let the plates go due to life. I'll do another round of transfers on the tea agar next. Maybe LI The next batch. Reading back on this thread, I was planning on transferring at day 3 before I got busy. The growth looked good but was slow. I'm hoping I cam peak at it soon and sneak in some transfers. Doubtful though.


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Edited by eatyualive (07/17/17 07:48 PM)


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Re: Cleaning dirty cultures with tea agar [Re: eatyualive]
    #24491224 - 07/18/17 08:50 AM (6 years, 6 months ago)

Great thread. Will definitely try this out. Thanks !


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Re: Cleaning dirty cultures with tea agar [Re: Snazz]
    #24495759 - 07/20/17 06:51 AM (6 years, 6 months ago)

The tea extracts are very acidic, did you check the end pH?


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24499609 - 07/21/17 08:19 PM (6 years, 6 months ago)

No I did not.
I'm assuming that is the reason the myc hasn't let off the wedge onto the plate yet? :lol:

Unless that normal for 4 days after transfer? I know I should expect slow growth but this is getting rediculous


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Re: Cleaning dirty cultures with tea agar [Re: ComebackKid]
    #24500333 - 07/22/17 07:38 AM (6 years, 6 months ago)

Here is C-Gel agar, which is another recipe I am trying, so far it works as intended, growth is quicker.
However I have no idea how well the C-Gel recipe will work for cubensis at this time.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24500382 - 07/22/17 08:24 AM (6 years, 6 months ago)

All my plates colonized except the culture of matias Romero I was cleaning up. Its about 50%.


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Re: Cleaning dirty cultures with tea agar [Re: eatyualive] * 1
    #24500922 - 07/22/17 02:19 PM (6 years, 6 months ago)

here is the matias romero. i believe this is after 7 days.



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Re: Cleaning dirty cultures with tea agar [Re: eatyualive]
    #24502183 - 07/23/17 06:16 AM (6 years, 6 months ago)

Very thick, and I see lots of variation. 12 and 6 o'clock look fast.

:thumbup:


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24502201 - 07/23/17 06:33 AM (6 years, 6 months ago)

If you are interested, add some CaCO3 powder until pH 7-8, which should increase speed.
The CaCO3 will also add more soluble carbon variation to the agar media.

Tea is acidic, as a bonus more of it breakdowns to CaCO3.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24502223 - 07/23/17 06:59 AM (6 years, 6 months ago)

Would there be any disadvantage to experimenting with tea in substrates? Just to cut down the chances of bacterial contamination during colonization in jars and spawn bags. Let's say a low dose of tea in the cooking water for grains? Moisturizing manure, straw, coir, ect using a low dose of tea-water instead of plain water? It sounds interesting. Never knew tea is such an effective anti bacterial agent.

Is it known why tea is so effective against bacteria? Is it just too acidic or???


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24502225 - 07/23/17 07:01 AM (6 years, 6 months ago)

Yeah plan on transferring when the fam leaves


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Re: Cleaning dirty cultures with tea agar [Re: eatyualive]
    #24502245 - 07/23/17 07:21 AM (6 years, 6 months ago)

Quote:

Shroomway said:
Would there be any disadvantage to experimenting with tea in substrates? Just to cut down the chances of bacterial contamination during colonization in jars and spawn bags. Let's say a low dose of tea in the cooking water for grains? Moisturizing manure, straw, coir, ect using a low dose of tea-water instead of plain water? It sounds interesting. Never knew tea is such an effective anti bacterial agent.

Is it known why tea is so effective against bacteria? Is it just too acidic or???




Here is a link to some natural alternatives, you can google more on tea extracts. Adding it to grain requires many steps.
You will need to correct the carbon-to-nitrogen ratio, and pH, which means testing, not sure it's worth it.

I did one test, the result was the bacteria did stall, but mold (mycelium) grew everywhere.
Ultimately it depends what contamination you get most often, mold or bacteria.



Quote:

eatyualive said:
Yeah plan on transferring when the fam leaves




Awesome, great idea with the media variation, looks like it's doing what you want.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather] * 1
    #24502480 - 07/23/17 09:51 AM (6 years, 6 months ago)

transferred those two sectors. one to another tea plate and the other to a pda plate. didn't waste time did the transfer before the fam was gonme 30 minutes. :rockon:


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Re: Cleaning dirty cultures with tea agar [Re: eatyualive]
    #24504474 - 07/24/17 07:30 AM (6 years, 6 months ago)

Has anyone tried my sucrose (carbon source) + soluble nutrients recipe, vs, malt extract agar?
It works really well for oysters, and produced strong growth, it's mildly antibacterial.


100g > CaCO3 Water, pH 7.5 - 8.2.

0.1g > Miracle-Gro Nutrients.
0.2g > 240B Gelatin.
2g > Sucrose.

2g > Agar


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24504520 - 07/24/17 08:10 AM (6 years, 6 months ago)





Gelatin (peptides): Composition.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24506809 - 07/25/17 09:41 AM (6 years, 6 months ago)

Here are the results of the C-Gel agar, and how to use it. And the next T-Gel agar recipe.
Thanks to, katbusa, Mycolorado and eatyualive for their test's and recipes.

You can also use it with malt extract, sucrose and so on.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24513689 - 07/28/17 09:51 AM (6 years, 6 months ago)

Here are the results of the updated T-Gel recipe, twice as strong, twice as fast.
Sorry for the image quality, there is condensate on the lid currently.

Scrapings also grow out, but growth is best from the peg.

   


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24514072 - 07/28/17 01:48 PM (6 years, 6 months ago)

Another comparison is, there is instant regeneration and growth on the peg, whereas recipe 1 did not regenerate.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24514084 - 07/28/17 01:50 PM (6 years, 6 months ago)

No starch, sugar or cellulose is being used in either recipe.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24514101 - 07/28/17 02:06 PM (6 years, 6 months ago)

Use the damn edit button


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Re: Cleaning dirty cultures with tea agar [Re: bodhisatta]
    #24514117 - 07/28/17 02:19 PM (6 years, 6 months ago)

Lol, sorry. Will do.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24525836 - 08/02/17 08:07 PM (6 years, 5 months ago)

Just popped by to show you my progress so far.
Ape myc on T agar
day 16


Took a while for it to leap off the wedge but after it did it seemed to be growing steadily.


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Re: Cleaning dirty cultures with tea agar [Re: ComebackKid]
    #24527647 - 08/03/17 02:28 PM (6 years, 5 months ago)

Was that scum formation already there when it set? Also is this a dirty culture?
If that is scum, then id say the mycelium outpaced the contamination.

If it's not contaminated, then I see adaption to media.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24546170 - 08/11/17 01:48 PM (6 years, 5 months ago)

So, what's the least amount of tea necessary to get benefits? Does PCing reduce the antibacterial properties? Could I just add a small pinch of lime, because I don't have a pH meter?

Also, from what I gather, tea can be mixed directly with agar, without adding nutrients from another source, is that correct? So, I could just make the tea, and add the agar powder, and voila?

I'm doing this right now. I used like 12 tea bags for maybe a gallon and a half of tea. I was just making some sweat tea for the family. It seems as though I probably didn't use enough tea, but I'll give it a go. I am now boiling the tea with some grains to add a bit of starch, and then I intend to add 50% H2O to that. Not as exact or sciency as some of you folks :wink: lol


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Re: Cleaning dirty cultures with tea agar [Re: Bigbadwooof]
    #24546717 - 08/11/17 05:47 PM (6 years, 5 months ago)

You can use 100% tea extract and agar for wood-loving, can't say for other mycelium.
If you know your mycelium can produce laccase then try 100% tea extract.

6-8g of lime to 100g of tea extract should be close.

Yes you can pressure cook it.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24546755 - 08/11/17 06:06 PM (6 years, 5 months ago)

But what concentration of tea? Normal drinking strength?

I believe I read one poster steeped 25g in a quart worth of water. It seems quite excessive to me. Then I read your post, which was I think 6g to 250ml (about the same)? What is the effective range. I assume it's preferable to stay on the lower end of that range, and I'd rather drink my tea :wink:


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Edited by Bigbadwooof (08/11/17 06:07 PM)


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Re: Cleaning dirty cultures with tea agar [Re: Bigbadwooof] * 1
    #24547646 - 08/12/17 04:22 AM (6 years, 5 months ago)

thanks for sharing the recipe for tea agar.
im gonna try this soon isolating from bacteria before doing slants.
since i cant source gentamicin from our area


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Re: Cleaning dirty cultures with tea agar [Re: Bigbadwooof]
    #24547669 - 08/12/17 05:03 AM (6 years, 5 months ago)

Quote:

Bigbadwooof said:
But what concentration of tea? Normal drinking strength?

I believe I read one poster steeped 25g in a quart worth of water. It seems quite excessive to me. Then I read your post, which was I think 6g to 250ml (about the same)? What is the effective range. I assume it's preferable to stay on the lower end of that range, and I'd rather drink my tea :wink:




I posted two recipe's for the T-Gel agar, one at 100% one at 50%, I am also using very cheap tea (60p for 250g).

"Extract: 3g (1 bag) Black tea + 125g CaCO3 water, check the end pH." -- So 1 bag to 125g water.


@pacmanbreed, Good luck m8.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24547830 - 08/12/17 08:31 AM (6 years, 5 months ago)

Quote:

Ferather said:
Quote:

Bigbadwooof said:
But what concentration of tea? Normal drinking strength?

I believe I read one poster steeped 25g in a quart worth of water. It seems quite excessive to me. Then I read your post, which was I think 6g to 250ml (about the same)? What is the effective range. I assume it's preferable to stay on the lower end of that range, and I'd rather drink my tea :wink:




I posted two recipe's for the T-Gel agar, one at 100% one at 50%, I am also using very cheap tea (60p for 250g).

"Extract: 3g (1 bag) Black tea + 125g CaCO3 water, check the end pH." -- So 1 bag to 125g water.


@pacmanbreed, Good luck m8.




CaCO3 = Soda water?

What if I just used tap water? I never checked the end pH, just hoping for the best. It solidified alright, as far as I can tell.

Sorry to bother you with so many questions. I ran out of gent, and this seems to be a preferable alternative anyway. Just want to make sure I'm understanding everything correctly, before I start wasting plates.

Thanks for the ratio of tea:water, btw.


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Re: Cleaning dirty cultures with tea agar [Re: Bigbadwooof]
    #24548634 - 08/12/17 04:19 PM (6 years, 5 months ago)

No "Ca" means calcium, not sodium. Too much sodium will kill or inhibit mycelium.
Other names: Chalk, Limestone, Oyster shell, Egg shell, Agricultural lime.

Ideally you want powdered forms, you can use a shaker on grit.



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Re: Cleaning dirty cultures with tea agar [Re: Ferather] * 1
    #24548764 - 08/12/17 05:31 PM (6 years, 5 months ago)

Oh, I have a bag of agricultural lime, and oyster shells. Sounds good!


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Re: Cleaning dirty cultures with tea agar [Re: Bigbadwooof]
    #24580409 - 08/26/17 05:14 AM (6 years, 5 months ago)

Gathered my ingredients today and want to try this for cleanin my 1st cube culture asap from bac.
Thank you for sharin ur effort and deep research brother ferather.

1.)[Q]Can I use pcied dogfood agar with added tea for cube cleanin?
We cant source LME from our area.
I do understand that pure tea will work on wood lovers that produce laccase for lingin.
Im still diggin on it. Tryin to Absorb good info from good fellow to also pass it someday.

2.)[Q]being dung lovers, can cube mycelium climb up in toothpicks to lessen my vectors from scalpel?
And is it advisable vs wedge on cubes when isolating sectors?

Hope you dont mind my noobish [Q].



Edited by pacmanbreed (08/26/17 05:33 AM)


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Re: Cleaning dirty cultures with tea agar [Re: pacmanbreed]
    #24580467 - 08/26/17 06:21 AM (6 years, 5 months ago)

1) I cant see why not, others are using standard recipe's for cubes and adding tea, with success.
2) Cubensis mycelium will happily populate a wood toothpick, for transfer or isolation.

The wood toothpicks will absorb, uptake soluble nutrients from the agar.


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Edited by Ferather (08/26/17 06:44 AM)


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24580573 - 08/26/17 08:00 AM (6 years, 5 months ago)

Love ur deep researched agar recipes brother.
Currently pcing them asm.
Will gladly report with pics in few days.


Been readin post lately. And found out this few minutes ago:
Quote:

Ferather said:
The peg will also receive nutrients from all through the agar, depending on the height of the peg.
This is due to evaporation, capillary action and mycelial uptake (distribution).


still learning more of forum  function. My bad.
Glad u have patience answering repetetive questions to help the new learners.:thanx:


Also found ur c-gel recipe.
Quote:

Ferather said:
The C-Gel agar is working, faster than T-Gel agar, and about the same speed as malt extract agar.
Both the MG solubles as nutrients and CaCO3 as carbon, are working as I intended.

Growth is slightly thicker than T-Gel, however still radial as normal.

   


Amazed how it produced nice rizho strands. :ohwow:
love to try it after sourcing MG alternative and 240bgel.
Love to see this on your sig brother someday.
Will try this on  dung lovers if time permits it.

- Can refined white table sugar be the source of sucrose?


Edited by pacmanbreed (08/26/17 01:35 PM)


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Re: Cleaning dirty cultures with tea agar [Re: pacmanbreed]
    #24580596 - 08/26/17 08:16 AM (6 years, 5 months ago)

This is great! With fall right around the corner, I can clone some of the local oyster without having to do 1000 transfers. It doesn't have to be pc'd, but I should be able to throw them in there with the rest of my no pours, and just keep them on hand for when they are needed right?


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Re: Cleaning dirty cultures with tea agar [Re: Mycolorado]
    #24580713 - 08/26/17 09:19 AM (6 years, 5 months ago)

Quote:

Mycolorado said:

200ml H2O
4.25g black tea (2 bags)
2g ME
4g Agar




Thanks for the recipe.
Gonna make 2 batches One with less nutrients(ME/DF).
And 50/50 tea from eatyou.


Edited by pacmanbreed (08/26/17 01:13 PM)


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Re: Cleaning dirty cultures with tea agar [Re: pacmanbreed]
    #24581338 - 08/26/17 01:54 PM (6 years, 5 months ago)

Seems to be working quite well on a bacterial culture I've had sitting on a slant for a long time. I have finally gotten it to fuzz up, instead of oozing like drooly ice cream. It's been on that slant for a couple years too, so it's about damn time I revive it! Gentamycin wasn't even working, btw.

In fact, the fact that this prevents spore germination makes it better than gentamycin in my book. As long as I use an intermediary MEA plate before going to grain, which I always do anyway, it will help keep things clean!


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Edited by Bigbadwooof (08/26/17 01:55 PM)


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Re: Cleaning dirty cultures with tea agar [Re: Bigbadwooof]
    #24581616 - 08/26/17 04:42 PM (6 years, 5 months ago)

Nice feedback.

:thumbup:


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24583234 - 08/27/17 12:50 PM (6 years, 5 months ago)

Made my 2 batches 2x7.
Both are 50/50 with varying dogfood amount.

- Hope this works as 240B gelatin substitue.
Having doubt on its sugars, carbs and sodium content.
I should have excluded it 1st, since DF is protien.

Will also try it on c-gel to verify them on second transfer without DF.


(A): 50g Water, 50g Extract, 2.5g Agar, 0.2g Gelatin, 0.75g Dogfood.
(B): 50g Water, 50g Extract, 2.5g Agar, 0.2g Gelatin, 0.50g Dogfood.


Edited by pacmanbreed (08/27/17 01:00 PM)


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Re: Cleaning dirty cultures with tea agar [Re: pacmanbreed]
    #24583431 - 08/27/17 02:29 PM (6 years, 5 months ago)

:takingnotes:


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24583678 - 08/27/17 05:19 PM (6 years, 5 months ago)

Quote:

Ferather said:
Nice feedback.

:thumbup:




Thanks :smile:

Thanks for doing the leg work on this one too! It has probably saved me a bit of headache, and $$$ (for antibiotics).


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24595241 - 09/01/17 06:46 AM (6 years, 4 months ago)

Quote:

Ferather said:
Has anyone tried my sucrose (carbon source) + soluble nutrients recipe, vs, malt extract agar?
It works really well for oysters, and produced strong growth, it's mildly antibacterial.


100g > CaCO3 Water, pH 7.5 - 8.2.

0.1g > Miracle-Gro Nutrients.
0.2g > 240B Gelatin.
2g > Sucrose.

2g > Agar




Just finished Cooking my shell grit water(PH 7.5)
It Will be used for c-gel.

Planing to use cgel for second transfer of my knockup 14 tgel if they successfuly colonize.
Have used:
12 tgel for cubes.
2 tgel fo wild ganoderma clone.
(Gonna post a separate thread on it)



- Will this work for MG substitute for cgel brother?


Also Dont have sucrose from sporeworks as of the moment, but have dextrose.
Im thinkering of activating yeast with table sugar for (maltose) production based on limited reading.
- Which sugar does mycelium prefer?
- sucrose, D-glucose or maltose?
(Maybe for glycogen production)

Hope you dont mind brother ferather.
apprecite doin your hard work on this,
testing various concentration with fellow shroom brothers to give a leap off for the new comers..
Really sparked my curiosity and love of growing.. hope can pass some knowledge to the little ones someday.

I still Dont have wide knowledge on nutrient/energy utilization yet.


Edited by pacmanbreed (09/01/17 08:18 AM)


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Re: Cleaning dirty cultures with tea agar [Re: pacmanbreed]
    #24596515 - 09/01/17 04:45 PM (6 years, 4 months ago)

That's an excellent substitute for MG. In terms of which sugar does best, I have personally only tried a few.
Sucrose (table sugar, white sugar), is easy and cheap to get, dextrose monohydrate I found best.

Dextrose monohydrate, or brewers sugar, is a D-glucose. Found in home brewing.
You will also find yeast nutrient in the same section, adds vitamins.

   


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24597733 - 09/02/17 07:47 AM (6 years, 4 months ago)

A note with the DIY agar recipe, it was never intended to be anti-bacterial, it's simply an alternative method.
It can be used for learning purposes as well, I guarantee vigorous mold in 3 days, be fully aseptic.

The good news is, it germinates spores beautifully, due to the soluble nature.
You can also use the same recipe, without agar for liquid cultures.

It is no different, other than DIY, to ME or PD and so on.
100% sugar and water will eventually stall.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24599732 - 09/03/17 02:57 AM (6 years, 4 months ago)

I just transferred from the tea agar plate to a few MEA plates. I didn't see any bacterial contamination on the tea plate. The culture I am growing has been on a slant for years, and it was quite heavily contaminated with bacteria. I cut a piece of colonized wood from a popsicle stick I had in the master slant, and used it to inoculate the tea plate, which I have now used to inoculate the MEA plates.

It is worth noting that when I made the tea plate, I also made an MEA plate from the same slant. The MEA plate has two distinct drips of bacteria going all the way to the edge of the plate, like drooly ice cream, but the rest of the plate appears to have grown out alright, to my surprise. The tea plate had very faint, tomentose growth, while the MEA plate had rhizomorphic growth. You can tell the age of the culture, by the rhizomorphic growth (on the original MEA plate), as it does not look terribly vigorous. The rhizomes are sparsely spread.

My phone is broken, so I can't post pictures.

One thing that I have been doing to combat contamination, that I think a lot of people do improperly, is to transfer only a very tiny sliver of mycelium from the very outer edge of visible growth, on to the next plate. I often see people transferring large wedges, and while that may speed up colonization of the plate, it also increases the potential for transferring contaminates.

Anyways, I will keep you guys posted, if you are interested in my results. I intend to take the 3rd transfer plate and make an agar slurry to inoculate jars with. Maybe I am jumping the gun, but I am really anxious to get things going again. I also have an AA+ culture that is equally old as fuck, which is contaminated with mycogone. I saw the mycogone growing on the plate, and I am attempting to transfer away from it, but this shit seems to be very difficult to get rid of.

TL;DR: Just made transfers from tea plates to MEA. All looks good. Hoping that continues, and I will keep you all posted!

I'm getting that sort of giddy impatience I had when I first started growing, because I want to see my precious old cultures fruit once again!


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Edited by Bigbadwooof (09/03/17 03:01 AM)


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Re: Cleaning dirty cultures with tea agar [Re: Bigbadwooof]
    #24600121 - 09/03/17 09:43 AM (6 years, 4 months ago)

Awesome work, keep it coming.

:takingnotes:


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24610756 - 09/07/17 04:26 AM (6 years, 4 months ago)

Cubes exhibits tementose on growth on t-gel.
This are 6 days from innoc. About half inch from the center.

bacterial donor brf plate


1st tgel cube wedge


2nd tgel cube wedge.

I think some spores has a hard/delayed time germinating on t-gel specially on high PH. Since Bacteria lowering the ph gives a window for eg. "green machine spores" to germinate.
Also when combating piggy backing bacteria from it I think that its better to control condensation and water pooling to maximize its effect and use toothpicks to leave the bacterial spore behind.


wood loving species loves the tea so much.
Garnoderma after 6 days. About 1.5+ inch from innoc.

1st wedge


2nd wedge

Hope Its not too late to plant toothpicks on them.
I will be away from home for a month due to work.
-Is it advisable to leave them colonizing and just transfer the toothpicks to new plates after a long 1.5 month?


Edited by pacmanbreed (09/07/17 04:31 AM)


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Re: Cleaning dirty cultures with tea agar [Re: pacmanbreed]
    #24611081 - 09/07/17 08:57 AM (6 years, 4 months ago)

The cubensis version, is this without sugar? If without, its done quite well for a sugar free media.
Obviously not the best growth ever, but it seems to have helped clean the sample.

The woodloving is doing well due to laccase as discussed.

:takingnotes:


Usually I would add the pegs, allow them to colonize and then refrigerate.
It should be ok to add the pegs and leave them if you have to.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24612860 - 09/08/17 01:10 AM (6 years, 4 months ago)

Quote:

Ferather said:
The cubensis version, is this without sugar? If without, its done quite well for a sugar free media.
Obviously not the best growth ever, but it seems to have helped clean the sample.

The woodloving is doing well due to laccase as discussed.

:takingnotes:

Usually I would add the pegs, allow them to colonize and then refrigerate.
It should be ok to add the pegs and leave them if you have to.




Thanks for the reply brother.
Yes its sugar less.
(B.) 50g Water, 50g Extract, 2.5g Agar, 0.2g Gelatin, 0.50g Dogfood.
Those are opened sacrificial cube plates for picture and control.
"Some are about 0.75-1" growth. From Recipe (A.) 0.75g DF agar recipe i posted".

Just finished cooking my pegs ready for instalation before leaving my work for a month.
Ill transfer the pegs to higher nutrition t-gel nextround.
50g Water, 50g Extract, 2.5g Agar, 0.2g Gelatin, 1g sugar 1g Dogfood.


Edited by pacmanbreed (09/08/17 01:14 AM)


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Re: Cleaning dirty cultures with tea agar [Re: pacmanbreed] * 1
    #24613154 - 09/08/17 07:29 AM (6 years, 4 months ago)

Wow ok, and it's still growing after transfer 3, that is very interesting. There is also no starch in the tea extract, T-Gel.
Since the cubensis shouldn't be producing laccase, it's running off the proteins in gelatin and dog food.

Proteins are composed of C-H-O-N, for the mycelium that is like sugar with nitrogen attached.
Based on the fact that most sugar's would otherwise count as a carbon source.

Humans also generate energy from proteins and C-H-O based vitamins.
100% protein or vitamins will produce a bad C:N ratio.

Ideally used as an additive to 100% C-H-O.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather] * 1
    #24613215 - 09/08/17 08:21 AM (6 years, 4 months ago)

Here is the composition of malt extract, and how to make an antibacterial agar recipe.
Copy the macro-micro nutrients, add a carbon source your mycelium likes.

Ideally the carbon source should be disliked by competitors.
This will be much harder for cubensis, no laccase.

Here is tea leaves, and tea extract.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24614658 - 09/08/17 08:28 PM (6 years, 4 months ago)

Quote:

Ferather said:
Here is the composition of malt extract, and how to make an antibacterial agar recipe.
Copy the macro-micro nutrients, add a carbon source your mycelium likes.
Ideally the carbon source should be disliked by competitors.
This will be much harder for cubensis, no laccase.
Here is tea leaves, and tea extract.



Pardon for my noobness.
Thank you for the links brother.
Im Thinkering of cellulose/other additive as C source, since coprophilic also digest this in nature.
:poopontoast:

Quote:

Ferather said:
Wow ok, and it's still growing after transfer 3



The Cubes are still on 1st transfer.

Had absorb that logical noob friendly explanation,
I think I made an out balance C:N ratio by using high protien DF & not adding sugar with tea as the only C source.
I think excess protien as N source is also attrative for competitors which i observed when ever i used DFA vs PDA based on mold germintion time leading to sooner sporulation..
Will source yeast extract next round for n and micro.
Ill be careful on N ratio nextime.


Based on your hard reseach, tea and gelatin is around 30:1 ratio.
Quote:


-what is outcome C:N ratio of my mix?
50g Water, 50g Extract, 2.5g Agar, 0.2g Gelatin, 0.50g Dogfood

-if youll use this on dung lovers, would you target it at 20:1 ratio?



.


Hope you dont mind my questions brother.
Ill try to use them as learning ground with trials.
Still absorbing computation of macro micro additives.
Eg. Homogenized HUMUS = 10:1.
Im still reading papers/links in regards to this.
Love how you used 50:1 ratio specially the pellets for wood lovers.


Edited by pacmanbreed (09/09/17 09:58 PM)


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24614956 - 09/08/17 10:42 PM (6 years, 4 months ago)

By the way an update before leaving home.
The leading edge of 1st t-gel cube sacrificial plate has grown to 1" and now visible after 2 days.
Also no signs of contam germination yet from opening it in a high spore load area. Will keep this control on how long till the landed green machine germinates on 7-7.5ph tea. They usually germinate in 3-5 days on DFA


1st tgel cube wedge comparison


Edited by pacmanbreed (09/09/17 10:02 PM)


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Re: Cleaning dirty cultures with tea agar [Re: pacmanbreed]
    #24616398 - 09/09/17 03:48 PM (6 years, 4 months ago)

My cultures look incredibly faint, and wispy on tea agar. They look like mold mycelium. Very tomentose. Maybe I put more tea in than you fellas.


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Re: Cleaning dirty cultures with tea agar [Re: Bigbadwooof]
    #24616589 - 09/09/17 05:24 PM (6 years, 4 months ago)

Quote:

Bigbadwooof said:
My cultures look incredibly faint, and wispy on tea agar. They look like mold mycelium. Very tomentose. Maybe I put more tea in than you fellas.



Did you add a Nitrogen source to this?
Mine look tememtose aswell. I guess its the tannins/tanic suppression nature of tea.

Ferather is right, I feel that we Just need to balance c:n ratios while using the teas macro/micro.
i feel that T-MEYA is standard starting/learning ground for this. But still can use any sources from our brother's pocket guide.


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Re: Cleaning dirty cultures with tea agar [Re: pacmanbreed]
    #24617478 - 09/10/17 03:40 AM (6 years, 4 months ago)

Malt Extract Yeast Agar?

I have actually considered adding a bit of coffee to it... I suppose then lime would become more of a necessity. I have some Bat Guano based marijuana fertilizer from fox farm... Maybe that would be a good nitrogen source.

As you can see, my approach to agar is much less scientific that some of you lol. In fact, the only time I measure my nutrient source, is when I'm using LME, because that shit is fucking expensive. Otherwise, I just use some watered down grain water, or something. I think I added grain juice to my tea agar for the first round I made. I don't remember.

Whatever I'm doing, it's working just fine, and I'm happy. I'm inoculating a bunch of jars now, with a beautiful looking clean culture. I think the bacteria had become resistant to gentamycin... I think I used gent on the master slant, years ago. Probably not the wisest decision lol!


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Re: Cleaning dirty cultures with tea agar [Re: Bigbadwooof]
    #24617670 - 09/10/17 07:46 AM (6 years, 4 months ago)

Quote:

Bigbadwooof said:
Malt Extract Yeast Agar?




yes just for observation purpose.
But we can exclude maltose and use other plenty of C sources around the kitchen.
Tea already has plenty of C for energy without starches and sugar but lacks in N.
Code:
Ferather said:
3g of sugar for every 100g tea leaves.
Potential energy and N sources (incomplete) for tea.
Polyphenols, varied: [ C6 | H5 | O1 ] x.
Cellulose, varied: [ C6 | H10 | O5 ] x.

Theophylline: C7 | H8 | N4 | O2.
Theobromine: C7 | H8 | N4 | O2.
Caffeine: C8 | H10 | N4 | O2.
Theaflavin: C29 | H24 | O12.
Tannins: C76 | H52 | O46.
Catechin: C15 | H14 | O6.
Carotene: C40 | H56.





Quote:

Bigbadwooof said:
I have actually considered adding a bit of coffee to it...




Minimal for N balance,
I feel that i will omit coffee since i always drink it  :drinkcoffee: , tea already has cafeine/sugar/macro/micro & its already acidic in nature around ph-4 with bacterial suppression properties due to tannins eg. "Camellia sinensis contain polyphenolic components with activity against a wide spectrum of microbes".
Ill use other N sources.
It seems thats cubes also love protien as N source.
Thats why gelatin&yeastextract is added to the mix.
Maybe its also the reason why it shows robust rizho in DFA due to protease production.



Quote:

Bigbadwooof said:
I suppose then lime would become more of a necessity.




Code:
RogerRabbit said:
mushroom metabolites will swing the pH acidic during colonization.

If you're using well water with a high pH over 8 or so,
a bit of vinegar or citric acid will lower it to neutral
- which is a good starting point.

I often say mycelium will colonize a bulk substrate fastest at around 5 to 6,
- to get people to avoid using lime which is recommended in so many teks.
for no reason because it often raises the pH too much, slowing colonization.

Most bulk substrate ingredients are already in the /- pH 6 range.
- so no adjustment is necessary.
RR



I feel this is the reason why our cubes exhibit very slow tementose growth in this agar mix..
They need to acidify the substrate 1st to around Ph of 5 - 6.5 to release carbon-acid bonds.
The Advantage of getting to ph of 7 is to increase suppressional properties and give a delayed/hard time for landed spores to release the sugars within after they germinate to prevent sporultion/muliply
Code:
Ferather said:
It works in the way of alternative carbon sources (non-sugar),
100g of black tea has 3g natural sugar no starch.
As it stands without the trace carbohydrates,
it's very nutritional and soluble, even cheap black tea.

The natural carbon containing acids breakdown at about ph 6.5, so pH 7-7.5 is ideal.
You can still get some bacteria but total activity should be reduced overall.
Here is some composition links (external),



Also Lime must be used in minimum amount just to balance out the acidic ph of tea.
Specially since some have high magnesium content based on sources. Unlike pure calcium oxide/CaO.
Better is to used egg shells in agar work to be safe, which is also a 1C source base on lewis structure.
Calcium carbonate: [Ca | C | O3]
Or 2c for Calcium bicarbonate: [ Ca | C | H | O3 ] 2.

Quote:

Bigbadwooof said:
I have some Bat Guano based marijuana fertilizer from fox farm... Maybe that would be a good nitrogen source.




Yes i think its good macro micro source aswell.
Better to use organic n source, i red from a paper that inorganic N reduces cellulase activty.
Code:
Ferather said:
Example nitrogen materials:
Proteins, Vitamins, Caffeine, Ureic, Ammonical.





Code:
Bigbadwooof said:
As you can see, my approach to agar
is much less scientific that some of you lol.
In fact, the only time I measure my nutrient source,
Is when I'm using LME,
because that shit is fucking expensive.


Same here brother.
Curiosity - community testing - observation- sharing - communication leads to greater knowledge & teks.

Glad elder ferather shared some of his leg work on this for community testings.
Im still gasping/absorbing to properly use our little stock in the kitchen.

I even cant source Lme from here. Importation is badly expensive and not an option for a kitchen grower.



Code:
Bigbadwooof said:
I think the bacteria had become resistant to gentamycin.
I think I used gent on the master slant, years ago.
Probably not the wisest decision lol!


Thank you for sharing ur observation.
Glad that saved me from start and some $$$$.


Hoping that elder brothers with greater understanding specially ferather shares a table of Potential energy and N sources of barley malt extract For easier comparsions to this. :smile:
Code:
Ferather said:
Potential energy and N sources (incomplete) for tea.

Polyphenols, varied: [ C6 | H5 | O1 ] x.
Cellulose, varied: [ C6 | H10 | O5 ] x.

Theophylline: C7 | H8 | N4 | O2.
Theobromine: C7 | H8 | N4 | O2.
Caffeine: C8 | H10 | N4 | O2.
Theaflavin: C29 | H24 | O12.
Tannins: C76 | H52 | O46.
Catechin: C15 | H14 | O6.
Carotene: C40 | H56.




Sorry for the long post. Ill use this for a reference while learning.


Edited by pacmanbreed (09/14/17 04:37 AM)


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Re: Cleaning dirty cultures with tea agar [Re: pacmanbreed] * 1
    #24617995 - 09/10/17 11:17 AM (6 years, 4 months ago)

So much input and good understanding. To clarify cubensis will mostly ignore the tea for carbon, but not the macro-micro nutrients.
Since the cubensis cannot digest the inhibitory materials, it will restrict the growth much like gentamicin or other.

For a woodlover, if not too much (too rich), it will digest the inhibitory materials, and grow pretty normal.
Those who have had good success using tea extract, used it like gentamicin, with normal agar.

Unless you are growing a woodlover, I'd take the route of treating it like gentamicin.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather] * 1
    #24618026 - 09/10/17 11:29 AM (6 years, 4 months ago)

I'm not 100% sure about the C:N that is best for cubensis, as far as I know 20:1 promotes fruiting, higher promotes colonization.
Since your using agar you probably want around 40:1 - 60:1 for a good speed. 100% grain is about 15:1 - 20:1.

2% nitrogen content total, 70% carbohydrates (44.45% carbon) = 31.115% carbon.

31:2 (31 /2 and 2 /2) = 15.55:1


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24618596 - 09/10/17 03:31 PM (6 years, 4 months ago)

Thanks for the guidance brother.
This will save me some precious time and efforts.
Since im goin to probio route & been composting for a year now.

Been drinking my coffee lately. :takingnotes:
Will limit it upon reading hydrate of carbons(carbohydate) glycogen etc.

Quote:

Ferather said:
2% nitrogen content total, 70% carbohydrates (44.45% carbon) = 31.115% carbon
31:2 (31 /2 and 2 /2) = 15.55:1



-(44.45% carbon) = average weight of C in most grain?

If so based on nutritional labels like this per 100g

-1.89% nitrogen content total, 72.8% carbohydrates (44.45% C weight) = 32.359% carbon
32.359:1.89(32.359/1.89 & 1.89/1.89) = 17.12:1

This is nice.
Im thinking of using the some of the sugar for probio ferment for my family to balance it to 30:1 before innoc.

Sorry for the bother and slightly being off topic.


Edited by pacmanbreed (09/11/17 08:18 AM)


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Re: Cleaning dirty cultures with tea agar [Re: pacmanbreed]
    #24622503 - 09/11/17 11:37 PM (6 years, 4 months ago)

Update:
11 days since innoc of 2 sacrificial plate outside SAb.
The one finally have contam germinated i can see sectoring at 11oclock.
The other one is almost fully colonized without visible contam.


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Re: Cleaning dirty cultures with tea agar [Re: pacmanbreed]
    #24622510 - 09/11/17 11:40 PM (6 years, 4 months ago)

why so much contam?


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Re: Cleaning dirty cultures with tea agar [Re: tombosley8]
    #24623976 - 09/12/17 04:43 PM (6 years, 4 months ago)

Quote:

tombosley8 said:
why so much contam?




They are sacrificial plates which are periodically opened in high spore load room to test the tea tannins effect on spore germination.

11 days sporulation is not quite bad.
I usually get contam sporulation around 3-4 days in dFA


Edited by pacmanbreed (09/12/17 04:44 PM)


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Re: Cleaning dirty cultures with tea agar [Re: pacmanbreed]
    #24625341 - 09/13/17 05:57 AM (6 years, 4 months ago)

Thanks for the informative post. You ought to post around here more often! :smile:


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Re: Cleaning dirty cultures with tea agar [Re: Bigbadwooof] * 1
    #24629978 - 09/14/17 07:22 PM (6 years, 4 months ago)

Yes it can happen, the green machine is a wood lover, and you are using its natural germination carbon :wink: .
That is a good test, 11 days of exposed situations, not bad. My mold test interestingly stalled.

The oyster mycelium overgrew it eventually, but that is wood lover vs wood lover.
Also the growth on the essentially protein agar seems usable, interesting.

Also your conversion of millet looks correct to me.

Composition of Gelatin (change to 100g).


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Edited by Ferather (09/14/17 07:42 PM)


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Re: Cleaning dirty cultures with tea agar [Re: Ferather] * 1
    #24630206 - 09/14/17 08:57 PM (6 years, 4 months ago)

Brown rice: 76% Carbs (inc fibre), 33.782% Carbon. 1.513 Nitrogen (usable).

33.782 / 1.513 = 22.32:1, Quite optimal for fruiting, faster in ratio, do 50/50 dry to dry cellulose-other to achieve: 44.64:1 (faster).

Starch and cellulose (plant fibre) are composed of the same components (sugars).


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Re: Cleaning dirty cultures with tea agar [Re: Ferather] * 1
    #24638782 - 09/17/17 05:21 PM (6 years, 4 months ago)

Quote:

Ferather said:
Yes it can happen, the green machine is a wood lover, and you are using its natural germination carbon :wink: .
That is a good test, 11 days of exposed situations, not bad. My mold test interestingly stalled.

The oyster mycelium overgrew it eventually, but that is wood lover vs wood lover.
Also the growth on the essentially protein agar seems usable, interesting.

Also your conversion of millet looks correct to me.

Composition of Gelatin (change to 100g).




The cubes also overgrew the green. I think its because of the ph of tea. (I feel that its Aspergillus from DF/corn vs trichoderma based on looks since i also keep contaminated plates for reference)
Interestingly my bacterial problem minimized and i can see sectoring clearly while the sectors grew flat on agar..
From this:

To this:


Good info, The gelatin has plenty of aminos and N for them. I think fast-carbon/sugar is what i lack from this recipe to give them a boost for transfers.
(B.) 50g Water, 50g Extract, 2.5g Agar, 0.2g Gelatin,
0.50g Dogfood.
Ill omit dogfood/starch and replace it with sugar next round.
I guess im getting the contam on the df high population load.
My bad last cooked plate didnt solidify, i forgot to balance the ph by cao3..

I think simpe tea+cao3+yeastnutrient+sugar is a good combo for macro+micro..


Edited by pacmanbreed (09/17/17 05:35 PM)


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Re: Cleaning dirty cultures with tea agar [Re: pacmanbreed]
    #24641101 - 09/18/17 02:59 PM (6 years, 4 months ago)

Thank you, I am currently trying another run from peg, using 2g of gelatin, since it also counts as a carbon source.
The only issue is the mixture needs more agar than normal, the media set very soft, its more like jelly.

The good news is, it does not liquefy when being digested, and appears to become harder.

I am tempted to rip open the tea bag and add directly to the agar powder.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24641176 - 09/18/17 03:27 PM (6 years, 4 months ago)

On a separate note, the cellulose agar alternative can produce some very nice genetics, essentially isolating growth on end media.
For a quick recipe, where used like agar for body, 25g dry paper pellets and 2-3g malt extract (or other agar additives).

You can cut down on the water content, as agar is around 95% water, ideal for attracting unwelcome germinating.
For example, 50% water, that's about 25g water added, 66% = 50g added, 75% = 75g added, and so on.

Use the weight system, as ratios of both water and nutrients come in grams, not volume.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24643076 - 09/19/17 05:48 AM (6 years, 4 months ago)

Forgot to mention, I am using CaCO3 (Calcium carbonate) with my agar, end result is pH 6.5, roughly.
I only got one spot of mold after 8 days, which stalled, no bacteria or yeast whatsoever.

I ran a test with CaCO3 as the only carbon source, with added Ma-Mi nutrients.
Its about 25% the strength of sugar per gram as a carbon source.

   


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24643107 - 09/19/17 06:15 AM (6 years, 4 months ago)

Moving back to plain WL-Tek (enriched, no added carbon), you can also use tissue samples.
The mycelium will use stored energy as it's starter carbon, it takes about 3-5 days.

I have also done, with added tea, and wild tissue (Image 2).

   



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Edited by Ferather (09/19/17 06:40 AM)


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24643282 - 09/19/17 08:12 AM (6 years, 4 months ago)

Thanks brother.
Quote:

Ferather said:
On a separate note, the cellulose agar alternative can produce some very nice genetics, essentially isolating growth on end media.
For a quick recipe, where used like agar for body, 25g dry paper pellets and 2-3g malt extract (or other agar additives).




Nice just like cattitives.

Quote:

Ferather said:
You can cut down on the water content, as agar is around 95% water




Confirm it. I guess this is for pure paper pellets water holding capacity = 95%?
Eg. 25g pellet + 25g water = 50% water?

Sorry for bumping again brother.
Would love to try this as soon as  got home.
Would try pure cellulose or pupal casing.


Edited by pacmanbreed (09/19/17 08:21 AM)


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Re: Cleaning dirty cultures with tea agar [Re: pacmanbreed] * 1
    #24643980 - 09/19/17 02:14 PM (6 years, 4 months ago)

No I mean a normal agar recipe will be around 95% water content as an end media.
With the pellets you are able to select your content from a given range.

For example: 2g ME + 2g Agar is 4g dry + 100g water (96%).
Pellets you can do 50% water, 66%, 75% so on.

----

Content range:

x 1 Dry Substrate | 1 Part(s) | 50% Water.
x 2 Dry Substrate | 2 Part(s) | 66% Water.
x 3 Dry Substrate | 3 Part(s) | 75% Water.
x 4 Dry Substrate | 4 Part(s) | 80% Water.


Usually I would remove the water present in pellets, but this can be ignored @ 5g per 100g.

So let me convert that into a working range:

A) 25g dry, x1 = 25g water, 50%. 1/2 dry, 1/2 water.
B) 25g dry, x2 = 50g water, 66%. 1/3 dry, 2/3 water.
C) 25g dry, x3 = 75g water, 75%. 1/4 dry, 3/4 water.

A) 25g + 25g, 50g total weight.
B) 25g + 50g, 75g total weight.
C) 25g + 75g, 100g total weight.


Tip: If you weigh your containers-setup, you can measure after cook content.


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Edited by Ferather (09/19/17 02:48 PM)


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24651141 - 09/22/17 08:01 AM (6 years, 4 months ago)

The 2g 240B gelatin T-Gel is working on Tarragon oyster, I'm using an 8 month old peg transfer, it regenerated in 3 days.

So far no contamination, here's the interesting part, there's no sugar so no I'm not pressure cooking.
I am also essentially assembling the entire media open air, however covered for cooling.

So far all attempts have produced usable mycelium, and in one case stalled mold.
As mentioned the Tarragon mycelium quickly devoured the stalled mold.

While I don't suggest what I did, it does show the usability.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24651153 - 09/22/17 08:08 AM (6 years, 4 months ago)

Additional recipe:

> Extract: 3g (1 bag) Black tea + 125g CaCO3 water, check the end pH.

> Recipe (C): 100g Extract, 3-4g Agar, 2g 240B Gelatin.

----

Additional notes:

Black tea, is normal green tea that has been oxidized, when tea oxidizes it turns very black.
When mycelium oxidizes tea, it also turn's it black, and you will see a dark ring.

Black tea is oxidized and easier to decay, as if done by laccase.
The good news is decay can occur without laccase.

Decay will also occur above pH 6.5.

----

Inhibitory materials present in plants are acidic, hence the fact most plants are acidic.
If these materials where not acidic or tolerant, they would decay (useless).

Acidic materials will happily breakdown via pH into carbon-other.

----

In short there are other natural methods of decay.

---

Side note: Cubensis is primary to cellulose, and requires many Ma-Mi nutrients.
In nature cubensis likes the nitrogen present in dung (quick-easy).

No laccase means it will have a tough time on some materials.
Especially if that material is void of fast carbon.

----

Breaking down a viable carbon source via pH using another viable carbon source = clever!


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Edited by Ferather (09/23/17 09:26 AM)


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24661404 - 09/26/17 12:34 PM (6 years, 4 months ago)

It seems 2g 240B gelatin is too much nitrogen, or there is not enough carbon in the tea extract.
Essentially the nitrogen ratio is far too high, growth is very good, but extremely slow.

I would therefore give the limit of around 0.8g 240B gelatin per 100g water.
If more is used, additional usable carbon will need to be added.

I'm going to try again, but using all of the tea, from dry.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24661523 - 09/26/17 01:03 PM (6 years, 4 months ago)

Quote:

Ferather said:
It seems 2g 240B gelatin is too much nitrogen, or there is not enough carbon in the tea extract.
Essentially the nitrogen ratio is far too high, growth is very good, but extremely slow.

I would therefore give the limit of around 0.8g 240B gelatin per 100g water.
If more is used, additional usable carbon will need to be added.

I'm going to try again, but using all of the tea, from dry.




I have to ask, why are you guys so interested in using gelatin instead of agar?


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Re: Cleaning dirty cultures with tea agar [Re: Bigbadwooof]
    #24661936 - 09/26/17 03:47 PM (6 years, 4 months ago)

It's not instead of agar, its being used as an additive along with the agar powder.
This is because mycelium will break it down for carbon and nitrogen.

Using just gelatin as the body, will turn to a liquid.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24666382 - 09/28/17 08:44 AM (6 years, 4 months ago)

Here is something I might try with T-Gel instead of calcium carbonate water, I can use generic indigestion tablets.
Cheap and ideal to use for agar, each tablet contains 500mg calcium carbonate, 524mg glucose.

The calcium carbonate will neutralize the pH (acidity) and add more carbon.

       


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24678427 - 10/02/17 03:33 PM (6 years, 3 months ago)

Here is the 2g gelatin test, much slower due to high nitrogen ratio, as I mentioned.
Due to the slowness, the black laccase oxidization ring is much larger.

It has a sweet smell, I hope due to glycogen production?

     

     


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24680833 - 10/03/17 12:10 PM (6 years, 3 months ago)

Quote:

Ferather said:
Here is the 2g gelatin test, much slower due to high nitrogen ratio, as I mentioned.
Due to the slowness, the black laccase oxidization ring is much larger.

It has a sweet smell, I hope due to glycogen production?

     

     




Oxidation ring? I haven't been following this thing from the beginning.


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Re: Cleaning dirty cultures with tea agar [Re: Bigbadwooof]
    #24681045 - 10/03/17 01:20 PM (6 years, 3 months ago)

Laccases (EC 1.10.3.2) are copper-containing oxidase enzymes found in many plants, fungi, and microorganisms.
Laccases act on phenols and similar molecules, performing one-electron oxidations, which remain poorly defined.

... Other laccases produced by the fungus Pleurotus ostreatus, play a role in the degradation of lignin ...

... Laccases require oxygen as a second substrate for their enzymatic action ...

Source.


--------------------
                   

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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24681055 - 10/03/17 01:22 PM (6 years, 3 months ago)

Black tea is a type of tea that is more oxidized than oolong, green and white teas.
Black tea is generally stronger in flavor than the less oxidized teas.

Source.


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Growing mushrooms, general guide and information (Ferather's Journal), https://ibb.co/rG3rML2

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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24704998 - 10/12/17 02:59 PM (6 years, 3 months ago)

Well the sweet smell as disappeared along with the oxidization ring, no signs of any contamination.
So in conclusion, the laccase and mycelium was converting materials into simple sugars.

I will add some wood pegs, and transfer some growth, takes 2-5 days.

Previous image, with laccase oxidization, and currently.

   


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24707034 - 10/13/17 12:15 PM (6 years, 3 months ago)

Next test recipe, cellulose agar alternative:

----

pH 7.5-8, No sugar and no starch.

0.1g > YN, soluble nutrients.
1g > MG, soluble nutrients.
75g > Boiling hot water.
25g > Paper pellets.
2g > 240B gelatin.
2-4g > CaCO3.

About 16:1 carbon to nitrogen.

----

Inoculated with a Tarragon peg: 16/10/2017 | 20:42 | UK format.

----


Basic nutrients in paper pellets:

Potassium, Phosphorus, Magnesium, Chromium, Copper, Iron, Manganese, Silicon.

----

Soluble nutrients provided by Miracle-Gro and Yeast nutrient:

   

   

----

Gelatin nutritional composition

----

Carbon sources:

Cellulose, Hemicellulose.
Calcium carbonate.
Proteins.

----

17/10/2017 | 14:30 | UK format.

Regeneration after about 12 hours, some growth.

   


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Growing mushrooms, general guide and information (Ferather's Journal), https://ibb.co/rG3rML2

https://www.shroomery.org/forums/showflat.php/Number/27857366#27857366

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Edited by Ferather (10/17/17 07:31 AM)


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #24717835 - 10/17/17 07:20 PM (6 years, 3 months ago)

I wonder if gentamycin+tea agar would slow growth too much.


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Re: Cleaning dirty cultures with tea agar [Re: Bigbadwooof]
    #24718796 - 10/18/17 07:10 AM (6 years, 3 months ago)

Quote:

Bigbadwooof said:
I wonder if gentamycin+tea agar would slow growth too much.




Slow doesn't mean bad, that also means it's working, or at least doing something to restrict growth.
I can get amazing genetics after leaving mycelium on a complex substrate (takes 4 months).

----

Custer's Chalk Medium (CaCO3 agar), used to identify growth via transparency.

----

Ferather's CaCO3 Agar:

The sucrose provides glucose and fructose, the soluble nutrients provides nitrogen and other nutrients.
Additional proteins, amino acids and other essential nutrients are also provided by the gelatin.


100g > Boiling Hot Water.

2-4g > CaCO3, Calcium Carbonate.
0.1g > MG, Soluble Nutrients.
2g > Sucrose (White Sugar).
0.2g > 240B Gelatin.
2.8g > Agar.

Assemble, microwave for 40 seconds.


Firmness: Harder than normal agar.
Colour: Cream-blue, opaque.





----

Calcium carbonate is white-off white, calcium bicarbonate is 100% clear.
Mycelium expel CO2, and acids, both react with CaCO3.


Calcium carbonate (insoluble): CaCO3 + CO2 + H2O → Ca(HCO3)2
Calcium hydroxide (insoluble): Ca(OH)2 + CO2 → CaCO3 + H2O

Calcium bicarbonate (soluble): Ca(HCO3)2


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Edited by Ferather (10/18/17 08:32 AM)


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Re: Cleaning dirty cultures with tea agar [Re: Mycolorado]
    #27210797 - 02/17/21 09:11 AM (2 years, 11 months ago)

Hey I'm a noob here.. Just started working with agar. I have some plates that wont stop transferring this gross slime with the mycelium, so I'm going to give this a shot, seems noob friendly. Hopefully it works I'll let you guys know.


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Re: Cleaning dirty cultures with tea agar [Re: infestedpasta]
    #27210848 - 02/17/21 09:36 AM (2 years, 11 months ago)

I had a slime contamination one time from vendor wood spawn, it tried to colonize but didn't do well (plain tea, no ME), I did have to transfer another time but all was well.
If you use a strong torch you can visibly see more contamination colonies than just the surface, making it easier to choose locations.

               

The bleaching (light orange) is white-rot, mycelial enzymes breaking down phenols.


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Re: Cleaning dirty cultures with tea agar [Re: Ferather]
    #27210930 - 02/17/21 10:31 AM (2 years, 11 months ago)

I just added some tea to grain water agar.. hopefully it works.


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