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Catattack some Agar 3.0

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#1 CatsAndBats

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Posted 25 January 2017 - 12:28 PM

Greetings again fellow amateur mycologists, autodidactic researchers and citizen scientists! Welcome to my new and improved agar-agar thread!

 

When I started my last agar thread it was set up as a learn as I go type thread, posting questions, problems etc. It also served as a log of sorts for recipes and my progress. This new thread is intended to be more of an active resource page. I will continue to log and display my work, but now that I have a couple of years working with agar, I feel that I  (along with my fellow "agar heads") can offer guidance and encouragement. Basically I'm cleaning up my other thread and placing all of the resources from it here in order to be accessed quickly and easily.

 

I get a lot of questions regarding homemade agar recipes and what type of agar should be purchased. Any powdered agar-agar from an Asian market will work. The agar strips are entirely too much work and aren't worth it IMHO. The basic recipe that I use for all of my own recipes is as follows:

 

10-12g agar powder (I've been using 12g but "standard" is 10g)

10g nutrition

600ml h2o (there's wiggle room here too 500-700ml works as well, just adjust the agar content accordingly)

stain (optional)

additives/future food (optional)

I refer to mine as "catattives" which are a pinch of all future substrates plus eggshells, pulverized and at the most add up to 0.5g total.

 

 

Now the above recipe can be adapted for virtually any purpose. If one were to use potato flake and corn sugar as the nutrition, one would have PDA agar, which is potato dextrose agar and is pretty universal in it's employ. One may use any sugar or "ose" as the nutrition, ie dextrose (corn sugar), sucrose (table sugar), maltose (yeast sugars), etc.. One may also use brown rice flour, dung, ground insects, and or any combination one may think of. I am of the opinion that if one has no previous agar  making experience, that one should stick to a basic carbohydrate/sugar source for the nutrition. At the end of this post I linked @seeker2be's aloha medicinal scholarship agar recipe index, which has many useful recipes and I use for reference.

 

It should be noted that activated carbon may be added as well to aid in spore germination, especially if the spores are older or their viability is questionable.

 

Here is an example of how I prepare a yeast agar that mycelium love:

 

 
post-147940-0-15627000-1468725370.jpg
 
The above is all of my ingredients that I'm playing with this particular round.
 
Included are 12g 'telephone' brand agar, a 7g packet of yeast , black food coloring, h2o, ~5g of my 'catattives' (powdered: eggshells, coir, oats, cornmeal, dirt, tiny bit of charcoal dust for good measure). 
 
In that picture there's a quart jar that has the activated yeast, which I activated by following the temperature parameters on the packet, which were 110-130f,  I stirred/shook until it was thoroughly consuming the 1/4tsp sucrose (table sugar) that I fed it, also on the packet. My goal was to get the yeast to produce maltose, which I hope coupled with my catattives, will be a terrific medium for my next set of transfers.
 
So basically after I got the yeast all riled up, I added water and everything together and shook vigorously, I then warmed it thoroughly on the stove, stirring a lot. I find this step seems to disperse whatever I've added as nutrition, and gets the agar thickened up, which helps keep all of my powders suspended, making my dishes uniformly nutritious. All together it was 700ml on the dot.
 
post-147940-0-43487700-1468725377.jpg
 
Here's the jars:
 

post-147940-0-60604700-1468725389.jpg
 

 
Here's the food coloring, yes I bought it because it had a black cat, go ahead, judge me. lol 
 

post-147940-0-73424700-1468725382.jpg
 
 
 

 

 

I did clean up this example so that it's only "meat and potatoes". It is just that, an example, and like my sample recipe it can be adjusted to one's particular needs.

 

I will update this tonight (hopefully) starting from where I left off on the former thread.

 

 

I would be remiss if I didn't reference and acknowledge my mentors and their agar threads. 

 

Uncle Microbe:

 

https://mycotopia.ne...3598-agar-work/

 

https://mycotopia.ne...crobes-randoms/

 

Peacefrog:

 

https://mycotopia.ne...le-and-recipes/

 

https://mycotopia.ne...mple-agar-prep/

 

Seeker:

 

https://mycotopia.ne...-log/?p=1198427

 

My boy whitethumb:

 

https://mycotopia.ne...-semi-grow-log/

 

Invisibility:

 

https://mycotopia.ne...nd-grow-thread/

 

And last but certainly not least, our beloved founder may he rest in peace:

 

https://mycotopia.ne...rs-agar-method/


Edited by catattack, 25 January 2017 - 12:44 PM.

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#2 MLBjammer

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Posted 25 January 2017 - 12:38 PM

Beautiful work, very well-organized and definitely educational. A lot of agar information is very scattered, so you are doing a heck of a service for many folks.

Thanks, my friend.
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#3 Arathu

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Posted 25 January 2017 - 12:41 PM

Out friggin standing! I'll be popping in on this one for sure.............. no judgements from me I like black cats.........

 

 

I refer to mine as "catattives" which are a pinch of all future substrates plus eggshells, pulverized and at the most add up to 0.5g total.

 

The fungi tell us if they like our mixes or not..........and then again I personally believe that introducing a fungi to the intended fruiting substrate "chemistry" early helps to select compatible strains as an environmental  default. 

 

Good stuff Cat!

 

I'm off tomorrow and plan on making a BRFHWA (brown rice flower hard wood agar) perhaps I shall get some black food coloring too............... :biggrin:

 

A


Edited by Arathu, 25 January 2017 - 12:43 PM.

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#4 CatsAndBats

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Posted 25 January 2017 - 12:49 PM

Out friggin standing! I'll be popping in on this one for sure.............. no judgements from me I like black cats.........

 

 

I refer to mine as "catattives" which are a pinch of all future substrates plus eggshells, pulverized and at the most add up to 0.5g total.

 

The fungi tell us if they like our mixes or not..........and then again I personally believe that introducing a fungi to the intended fruiting substrate "chemistry" early helps to select compatible strains as an environmental  default. 

 

Good stuff Cat!

 

I'm off tomorrow and plan on making a BRFHWA (brown rice flower hard wood agar) perhaps I shall get some black food coloring too............... :biggrin:

 

A

 

If you want to be extra baller, use carbon as the stain. Here's what 1g of carbon does:

 

post-147940-0-34490300-1478094987.jpg

 

I imagine 2g would do the trick with the added benefits from the carbon!


Edited by catattack, 25 January 2017 - 01:01 PM.

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#5 Arathu

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Posted 25 January 2017 - 01:02 PM

Beautiful stuff..........perhaps even carbon made from the target hardwood............hmmmm.........interesting. Wonder if the yellow morels would be into it? 


Edited by Arathu, 25 January 2017 - 01:02 PM.

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#6 Jeepster

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Posted 25 January 2017 - 01:16 PM

GREAT thread!!!
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#7 Nsnail

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Posted 25 January 2017 - 01:20 PM

You've done it again cat, great stuff.

Was thinking earlier about training with small amounts of poo


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#8 Microbe

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Posted 25 January 2017 - 01:45 PM

Very proud brother! I see you adopted the 'ose' nutes for explaining carbon sources!

This thread has to be archived material already as it stands. Mods what do you think? Hell yeah it is sais thr Mods ;)

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#9 CatsAndBats

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Posted 25 January 2017 - 01:47 PM

Very proud brother! I see you adopted the 'ose' nutes for explaining carbon sources!

This thread has to be archived material already as it stands. Mods what do you think? Hell yeah it is sais thr Mods ;)

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Let's get more knowledge on it first, thanks uncle microbe!


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#10 Microbe

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Posted 25 January 2017 - 01:51 PM

I have said this before but i dont think many know how much time is involved to lay down a structured thread like this. Your not getting paid to do this, your doing this out of your intense passion to teach and help evolve the cultivator of any skill set or tenor. It is very educational and easy to follow along. Perhaps now that i have nothing but time on my hands i may assist in these teachings. Again brother excellent work and thank your putting together a valuable resource for those who want to learn about the backbone of the hobby. However im deducting a few points and grading this a A-. You know why dont you? I bet you do......

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Edited by Microbe, 25 January 2017 - 01:52 PM.

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#11 CatsAndBats

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Posted 25 January 2017 - 03:27 PM

Hahaha! Petri dishes!
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#12 Ferather

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Posted 25 January 2017 - 04:14 PM

You can also beef up your agar by adding 1-2 teaspoons of 240B Strong Gelatin for extra peptides.


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#13 CatsAndBats

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Posted 25 January 2017 - 04:17 PM

You can also beef up your agar by adding 1-2 teaspoons of 240B Strong Gelatin for extra peptides.

 

Literally and figuratively "beef it up".

 

Well "pig it up" from the link. :biggrin:


Edited by catattack, 25 January 2017 - 04:18 PM.


#14 Ferather

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Posted 25 January 2017 - 04:21 PM

If you live in the UK, like me, you can also get: xanthan gum and guar gum.

As well as many other laboratory-food grade additives from here.

 

:biggrin:


Edited by Ferather, 25 January 2017 - 04:29 PM.


#15 archersmark

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Posted 25 January 2017 - 06:57 PM

I picked up some telephone brand agar per your earlier Agar tek.  The only thing I'm on the fence about using is food coloring.  When I did my contamination tests for mold and bacteria in my clean room last summer, I had a lot of black and dark green mold/bacteria that I worry wouldn't stand out on my agar.  This is me being anxious for sure.  I do want to do at least one round with black food coloring, just to see the beautiful white mycelium stretching and exploring across the surface in bold, vivid contrast to the black background of the agar.

Contaminants:

e.jpg

 

8.jpg


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#16 CatsAndBats

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Posted 25 January 2017 - 07:10 PM

I picked up some telephone brand agar per your earlier Agar tek.  The only thing I'm on the fence about using is food coloring.  When I did my contamination tests for mold and bacteria in my clean room last summer, I had a lot of black and dark green mold/bacteria that I worry wouldn't stand out on my agar.  This is me being anxious for sure.  I do want to do at least one round with black food coloring, just to see the beautiful white mycelium stretching and exploring across the surface in bold, vivid contrast to the black background of the agar.

Contaminants:

attachicon.gife.jpg

 

attachicon.gif8.jpg

 

Stain is optional as well as the color. I typically change the color on mine with every batch or use color to identify the type of agar that I made. For example when I started I'd use blue for 'spore germination' agar. Now I just pick a new color with each batch so when I look at my cabinet I can organize them in my head quickly.

 

The molds turn green or black when they sporulate and at that point the dish or the jar is screwed IMHO. Bacteria and competitor yeasts are easier to spot regardless of stain.



#17 peacefrog

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Posted 25 January 2017 - 07:15 PM

Very nice write up as always, cat! Thank you for putting this together. You da man.

#18 CatsAndBats

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Posted 25 January 2017 - 07:19 PM

 

post-147940-0-48767100-1479434773.jpg

Malabars continuing to look better. Nice radial growth.

I want to see it run the low nutrient agar I made up.

That is uniform growth and pretty dang close to a isolate. Dont let the lack of that roping rhizo fool you. That is considered rhizomorphic growth btw which you know that.

ecc4fed1d8a7d5608dcfecc4f422a0f4.jpg


I want to talk about this. Where the arrow is pointing is the filaments of hyphae branching out ahead of the mycelia colony. This is where the chemical signals are happening but what i really want to say is that is where you want to transfer from when trying to isolate a monokaryon but a little sooner and as soon as you see that haze. Obviously thats not a monokaryon but you will see the same thing from a monospore germination but you have to grab it as soon as you see the first sign and from the edge.

Now something else i have seen and never even thought of this, i have seen monokaryotic mycelium left behind. Where mating has taken place and it is now dikaryon growing out with the monokaryon left in the back field so to speak, you could grab it from there if that ever happened and with confidence. You would need to do it before the plate became colonized because the dikaryon will most certainly will start growing back over the monokaryon.

You know what im saying homeboy?

 

 

 

Above is a quote from me an uncle microbe discussing my malabar culture. The point of me placing this here is two fold. Firstly, if there was a competitor's mycelium (mold) or a bacterial/yeast contamination, one would be able to see it very clearly on the surface. Secondly, as microbe keenly points out, one can even see the chemical signals sent by the mycelium on the agar surface. Original here: https://mycotopia.ne...gain/?p=1295174

 

Thanks @peacefrog, just trying to make you proud!


Edited by catattack, 25 January 2017 - 07:21 PM.

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#19 Microbe

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Posted 25 January 2017 - 07:45 PM

Hahaha! Petri dishes!

Told you!

#20 Microbe

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Posted 25 January 2017 - 08:45 PM

I picked up some telephone brand agar per your earlier Agar tek. The only thing I'm on the fence about using is food coloring. When I did my contamination tests for mold and bacteria in my clean room last summer, I had a lot of black and dark green mold/bacteria that I worry wouldn't stand out on my agar. This is me being anxious for sure. I do want to do at least one round with black food coloring, just to see the beautiful white mycelium stretching and exploring across the surface in bold, vivid contrast to the black background of the agar.
Contaminants:
attachicon.gife.jpg

attachicon.gif8.jpg

I use it because that first sign of growth is very easily observed and identifiable in the very early stages. But here is something to think about for long term storage. Before i post the next paragraph let me say i have stored cultures for over a year on less then a 1/8" poured plate stored in a ziploc bag and the cultures were very viable. Imagine slants with 2" or more with the preservative benzoic acid, the preservative in most commercially produced food colorings. My myc loves it!

C&P from Wiki

The mechanism starts with the absorption of benzoic acid into the cell. If the intracellular pH falls to 5 or lower, the anaerobic fermentation of glucose through phosphofructokinase decreases sharply,[13]which inhibits the growth and survival of microorganisms that cause food spoilage.

Further reading....

https://en.m.wikiped...Sodium_benzoate

Edited by Microbe, 25 January 2017 - 08:47 PM.






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