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Peacefrog's agar style and recipes


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#41 CatsAndBats

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Posted 10 November 2015 - 04:04 PM

I might give that a try, but it's not so much the bacteria or competing fungi or molds, at least not lately since I changed over to jars.

 

It's just that the sample sits there forever and does nothing.

what are you using for the nutrition? 

 

Plus, for whatever culture that I'm trying to transfer, I make 5 jars minimum, hedging bets.


Edited by catattack, 10 November 2015 - 04:07 PM.


#42 happy4nic8r

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Posted 10 November 2015 - 06:08 PM

I make 1 liter, tap water

            20g    agar

            20g    malt dextrose

              1g    soy peptone   optional

              1g    yeast              optional

 

mix it with hot water in the blender and pour jars, petrie dishes, remainder into clear glass bottles

 

put in pc for 30 - 40 mins, when cold tighten jar lids, wrap parafilm around plates, cork bottles.

 

All go into the glove box, except bottles which go in fridge.

 

I have also used the bottles stored in fridge by microwaving until melted and hot, and poured more plates with that.

 

Sometimes it works. Not 100% works, but 50 - 75% will grow without issue.

 

Sometimes I have had it not harden at all, sometimes it hardens like a brick, these out of the same bottle.

 

It is such a random success thing with me that I really just stick to other things that work, but i still keep trying. 

 

I found that adding the ingredients slowly in the blender on slow with hot *(not boiling) water made a good clear mix.

 

Trying to boil it on the stove was time consuming, messy and had clumps that were hard to mix out.

 

Is there something I'm doing wrong here?



#43 peacefrog

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Posted 10 November 2015 - 06:38 PM

There seems to be nothing wrong with your recipe there. But there is really no need to blend in a blender. If you are using the full recipe in bottles and pouring into plates, all you need to do is add the water and the dry ingredients shake well and sterilize. If using jars, just add the water then a heaping teaspoon or so of dried agar, mix with a spoon and sterilize.

I wouldn't recommend microwaving agar. You really can't control the temps that way. You could be carmelizing the sugars/starch in there rendering them inaccessible for the mycelium to ingest.

The more agar agar you use, the stiffer the gelatin like state it will be. If you use too little, all should grow well, it just might be a little loose.

I think you are over thinking this. Like said above, add the dry ingredients to water cool or warm, and mix. It really doesn't matter the temps of your water at this stage. The sterilization will do the rest.
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#44 CatsAndBats

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Posted 10 November 2015 - 06:56 PM

cat edit, just saw @peacefrog's answer..

 

 

I just got my agar to work consistently, so I'm not experienced enough to spot problems but here's what I do:

1qt h2o

10g agar

10g <powdered nutrition, usually dog treats/food>

 in grinder, then simmer, back into quart to be shaken before each pour, lids, PC 20min 15psi cool as SLOW as possible to avoid excessive moisture.

 

I always use jars and @peacefrog taught me to flip them to prevent the excess moisture from hitting the agar.

 

 

@peace, do you see any flaws with mine?


Edited by catattack, 10 November 2015 - 07:02 PM.

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#45 CatsAndBats

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Posted 10 November 2015 - 08:36 PM

@peacefrog and community, do you think adding dead mold to agar would train the mycelium to have the correct enzymes to digest live mold? Make it's own antibiotics?


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#46 whitethumb

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Posted 10 November 2015 - 10:24 PM

First off congrats, cat!! Nice agar porn. And I agree with mikey, not quite an isolate yet, but you are on the right path, my freind. It could take several transfers sometimes to get a pure culture. All growth will be uniform no matter what type of mycelial growth you are trying to isolate. Here is an example of what mikey is talking about:
attachicon.gifimage.jpeg

And here is an example of an isolate of linear type mycelium:
attachicon.gifimage.jpeg

Happy,

I also must agree with mikey. I have kept pre-made agar for a few years as well with good results. MEA clumps up the most because of the malt. You really have to keep malt sealed from moisture. But once introduced to hot water, it dissolves and is fine. I have never attempted using antibacterial agar before, but it might be a good thing for you to try. What is your procedure with agar? Perhaps give us some insight and pics as well?

Kind of like this? ;)
uploadfromtaptalk1447212252483.jpg
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#47 CatsAndBats

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Posted 10 November 2015 - 10:59 PM

 

First off congrats, cat!! Nice agar porn. And I agree with mikey, not quite an isolate yet, but you are on the right path, my freind. It could take several transfers sometimes to get a pure culture. All growth will be uniform no matter what type of mycelial growth you are trying to isolate. Here is an example of what mikey is talking about:
attachicon.gifimage.jpeg

And here is an example of an isolate of linear type mycelium:
attachicon.gifimage.jpeg

Happy,

I also must agree with mikey. I have kept pre-made agar for a few years as well with good results. MEA clumps up the most because of the malt. You really have to keep malt sealed from moisture. But once introduced to hot water, it dissolves and is fine. I have never attempted using antibacterial agar before, but it might be a good thing for you to try. What is your procedure with agar? Perhaps give us some insight and pics as well?

Kind of like this? ;)
attachicon.gif2015-07-02 06.39.19_noexif.jpg

 

 

I'm almost there, I took pieces from the best jar and made 5 more, do I just look for the most aggressive and try to get one radial? 


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#48 whitethumb

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Posted 10 November 2015 - 11:31 PM

First off congrats, cat!! Nice agar porn. And I agree with mikey, not quite an isolate yet, but you are on the right path, my freind. It could take several transfers sometimes to get a pure culture. All growth will be uniform no matter what type of mycelial growth you are trying to isolate. Here is an example of what mikey is talking about:
attachicon.gifimage.jpeg

And here is an example of an isolate of linear type mycelium:
attachicon.gifimage.jpeg

Happy,

I also must agree with mikey. I have kept pre-made agar for a few years as well with good results. MEA clumps up the most because of the malt. You really have to keep malt sealed from moisture. But once introduced to hot water, it dissolves and is fine. I have never attempted using antibacterial agar before, but it might be a good thing for you to try. What is your procedure with agar? Perhaps give us some insight and pics as well?

Kind of like this? ;)
attachicon.gif2015-07-02 06.39.19_noexif.jpg

I'm almost there, I took pieces from the best jar and made 5 more, do I just look for the most aggressive and try to get one radial?
It depends on what your sole purpose is. If it's to get even uniform grow, i like to take a section that's fast growing and uniform.
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#49 coorsmikey

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Posted 11 November 2015 - 05:37 AM

@peacefrog and community, do you think adding dead mold to agar would train the mycelium to have the correct enzymes to digest live mold? Make it's own antibiotics?

The Mycelium already loves to eat dead mold and live mold for that matter, the problem is when other molds eat our mycelium. How do you train our myc to be stronger and less desirable to competition organisms? Food for though for ya?
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#50 CatsAndBats

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Posted 11 November 2015 - 08:54 AM

 

@peacefrog and community, do you think adding dead mold to agar would train the mycelium to have the correct enzymes to digest live mold? Make it's own antibiotics?

The Mycelium already loves to eat dead mold and live mold for that matter, the problem is when other molds eat our mycelium. How do you train our myc to be stronger and less desirable to competition organisms? Food for though for ya?

 

the agar really has my brain working!

 

@mikey my thought is to make a super agar. training the myc to already have the enzymes produced to eat most subs and contaminates it may encounter. 



#51 coorsmikey

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Posted 11 November 2015 - 01:15 PM

@catattack, once you get you agar isolation technique going and when you using domesticated strain from reputable growers, most myc does do what you are describing. Here are some pics of I think you are talking about. Although these are on there way to the garbage as I would not use them personally, I thought why not share so they are not a 100% waste.
These two are pics of ps.cyan over coming a nasty black mold.
image.jpeg image.jpeg
These two are a trich contaminated LC of SG-30 where you can see the myc feeding of off the trich.
image.jpeg image.jpeg
I have no intention of using these or trying to clean them up. Their final destination is the compost pile.

Edited by coorsmikey, 11 November 2015 - 01:15 PM.

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#52 peacefrog

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Posted 11 November 2015 - 08:07 PM

Cat,

Your recipe sounds fine. 1 would recommend using 20 grams of agar per quart of water, but like said before, it doesn't really matter how much agar you use. It's all about the consistency you are looking for and comfortable with. As for looking for mycelium that will overcome contamination, I am not the best person to ask. I have never attempted those kinds of experiments. I think mycrobe has, but like mikey, I either toss or attempt to clean up a contaminated plates.

My advice would be to transfer very tiny pieces of sectors that, like whitethumb says, are fast and uniform. Sometimes the best sectors you can find will not provide you with a uniform culture. You may have to transfer many times. The more plates you have, the better. Then once you have a few great looking mono cultures, you must test them. They all will produce but the best looking culture could either be great, good or shitty.

Edited by peacefrog, 11 November 2015 - 08:09 PM.

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#53 Heirloom

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Posted 14 November 2015 - 01:56 PM

Thank you peacefrog, this amounts to a guide, instructions. I will study this as my agar skill needs improving.
  I can't tell you how useful your discourse is.

 goog vibes


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#54 Lakegal7

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Posted 16 November 2015 - 10:44 PM

Peacefrog, thanks for sharing. I have been intimidated with working with agar, I will try your methods.
Thanks to all who contributed to this thread/forum
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#55 MLBjammer

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Posted 17 November 2015 - 04:38 AM

This approach to agar is great in that it removes the intimidation factor from working with agar.  Everybody has a few 1/2 pint jars and solid lids around the house, if they have ever grown cakes.


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#56 CatsAndBats

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Posted 17 November 2015 - 07:14 AM

This approach to agar is great in that it removes the intimidation factor from working with agar.  Everybody has a few 1/2 pint jars and solid lids around the house, if they have ever grown cakes.

 

Agreed! 

 

It's funny because I was gifted an aa+ with a big old mold spot and transferred until I had a clean culture. Now I'm staining own my agar recipes.. go figure!


Edited by catattack, 17 November 2015 - 07:14 AM.

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#57 maxintensity

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Posted 17 November 2015 - 07:46 AM

This thread is awesome! Thanks for all the great info! I'm incredibly excited to venture into agar. A big thanks to a fellow topia member for being incredibly generous sending me some plates of a few different edibles. Time to get rocking on this!
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#58 CatsAndBats

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Posted 17 November 2015 - 07:48 AM

This thread is awesome! Thanks for all the great info! I'm incredibly excited to venture into agar. A big thanks to a fellow topia member for being incredibly generous sending me some plates of a few different edibles. Time to get rocking on this!

creepy voice: join us..

 

hahaha!


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#59 Heirloom

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Posted 03 December 2015 - 02:45 PM

This helped me over come my fear , I did little work in the past, now I have cloned for the first time, nice growth. used a still air box and prepared agar bought.

 


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#60 Nsnail

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Posted 17 January 2017 - 11:15 PM

Yo I'm trying to use your recipe here and things aren't adding up. To clarify it's 5 grams of brf and 5 grams agar to 15ml of water? It comes out way too dry


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