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Nice Ol Bud
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Registered: 09/11/13
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Loc: Mystical Maze of Mushroom...
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Agar Formula Help
#19317237 - 12/23/13 05:15 PM (10 years, 1 month ago) |
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Well I just don't understand the ratios. I have Karo, Potato Flakes, & Agar available.
Some people say 5g potato flakes. Some say 20g's.
Can I just go with
10g's Agar 10g's Potato Flakes 10ml's Karo 500ml Tap Water.
Would this be sufficient? Any help from the more experience would be great.
And what's the purpose of Karo/Dextrose in the mix, scientifically. Also what role do potato flakes have in the mix?
Thanks guys!
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Psilicon
Really Nice Guy


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Knock the potato flakes down to five grams.
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Nice Ol Bud
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Re: Agar Formula Help [Re: Psilicon]
#19317284 - 12/23/13 05:24 PM (10 years, 1 month ago) |
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Why so little? Why do people recommend 20grams flakes and 15grams agar per 500ml of water?
I don't get it? Does the mix not really matter with agar prep?
So many different ratios.
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Psilicon
Really Nice Guy


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Oh, it matters very much. 10mL of karo or honey is roughly 7g of dextrose/glucose, and almost all of the potato is starch. Together that brings them to 2.4% available sugars, which is ideal for your purposes. With mushrooms, any less and they don't have enough nutrients to properly grow, and any more and they see no reason to. Too much sugar result in slow, fluffy, tomentose growth which is harder to isolate and more prone to contamination as a result.
2-3% is perfect for both agar and sugar concentration. Any more or less and you stray from the ideal.
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Nice Ol Bud
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Re: Agar Formula Help [Re: Psilicon]
#19317419 - 12/23/13 05:48 PM (10 years, 1 month ago) |
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Well that just explained a good amount to me. How'd you determine those percentages?
So your saying
5g Potato Flakes 10g Agar 10ml Karo 500ml Tap Water
So that's a perfect mix right their?
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Psilicon
Really Nice Guy


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Yep, all of those are within the ideal range. I personally like a stiffer agar, so I would take that as high as 13g agar agar for that quantity, but I know there are many talented people out there doing agar work who like it right at the 2% mark. I'd try 12g agar agar and adjust from there next time.
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twistedty
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Re: Agar Formula Help [Re: Psilicon]
#19317447 - 12/23/13 05:53 PM (10 years, 1 month ago) |
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as long as you get the consistency of the agar down you want it kinda gelled, myc will colonize mostly anything
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Nice Ol Bud
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Re: Agar Formula Help [Re: twistedty]
#19317493 - 12/23/13 06:03 PM (10 years, 1 month ago) |
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Alright great. Also where did you get that 10ml Karo is 7grams of dextrose? Also and what is starch? A sugar as well?
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Psilicon
Really Nice Guy


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Starch is a polysaccharide. It's a complex carbohydrate made up of lots and lots of simple sugars strung together in a sort of daisy chain of energy. Bacteria are pretty lousy at processing them, as are yeasts, but complex fungi can produce amylases, which are enzymes that break down starches into their component sugars. This gives our fungi a distinct advantage on agar.
You have amylase in your saliva, by the way. If you put something starchy in your mouth, like potatoes, popcorn or even wheat crackers, after a little while you'll start to notice a sweet taste. That's from the simple sugars that have been cleaved off the starch molecules, and which now fit into your taste bud receptor enzymes.
If you don't want to wait around with a pasty white powder in your mouth long enough for it to turn sweet (I don't blame you), just put a drop of iodine on it. If it turns blue-black or purple, you've got starch. If it stays red or orange, there's no starch.
So there you go. Now you know what starch is, why we use it, and how to identify it. Along with the ideal percentages for agar plates, that's all you need to make up your own agar recipes.
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Psilicon
Really Nice Guy


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Quote:
Nice Ol Bud said: How'd you determine those percentages?
It's just a matter of some really easy math and knowing the density of water. Fortunately, the beautiful thing about the metric system is that we use water so much that literally every unit of measurement is based on water, so here's something easy to remember:
1cm^3 (or cc or cubic centimeter) of water is 1mL. 1mL of water weighs 1g at room temperature. Every degree Celsius is 1% of the difference between boiling water and freezing water, and a calorie is how much energy you need to raise a gram(or cc or mL) of water one degree Celsius, but that's not really relevant. And despite this, people say still SAE is easier. 
So let's say you have 5g of starches (which, as you can see from my earlier post, is actually 5g of simple sugars strung together in chains). You also have 7g of dextrose. 5g + 7g is 12g. So what percentage of the total solution is sugars?
First we need to ask ourselves, "Roughly how much will a 500mL solution weigh?" Since every mL of water weighs a gram, the answer is obviously 500g water + 12g sugars, for a total of 512g. Then just a little simple math:
12g sugars = 2.34% sugars. 512g solution
Does that help?
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Stromrider
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Re: Agar Formula Help [Re: Psilicon]
#19322620 - 12/24/13 07:07 PM (10 years, 1 month ago) |
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I've lived in America my whole life and was raised with the sae system and I fucking hate it! I'll never understand why we haven't abandoned it completely. Love the metric system
That was some great info van. I appreciate that
Nice to see you ol bud. How ya been
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Lazarus.Long
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Registered: 09/10/13
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Re: Agar Formula Help [Re: Psilicon]
#19323062 - 12/24/13 08:21 PM (10 years, 1 month ago) |
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Quote:
van der griegen said:
Quote:
Nice Ol Bud said: How'd you determine those percentages?
It's just a matter of some really easy math and knowing the density of water. Fortunately, the beautiful thing about the metric system is that we use water so much that literally every unit of measurement is based on water, so here's something easy to remember:
1cm^3 (or cc or cubic centimeter) of water is 1mL. 1mL of water weighs 1g at room temperature. Every degree Celsius is 1% of the difference between boiling water and freezing water, and a calorie is how much energy you need to raise a gram(or cc or mL) of water one degree Celsius, but that's not really relevant. And despite this, people say still SAE is easier. 
So let's say you have 5g of starches (which, as you can see from my earlier post, is actually 5g of simple sugars strung together in chains). You also have 7g of dextrose. 5g + 7g is 12g. So what percentage of the total solution is sugars?
First we need to ask ourselves, "Roughly how much will a 500mL solution weigh?" Since every mL of water weighs a gram, the answer is obviously 500g water + 12g sugars, for a total of 512g. Then just a little simple math:
12g sugars = 2.34% sugars. 512g solution
Does that help?
I just gave you a +5 for the ease with which you explained this concept, Thank you sir!!!
Now many of the other Agar threads suddenly make much more sense.
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Gr13nMushDude
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Re: Agar Formula Help [Re: Psilicon]
#19323093 - 12/24/13 08:30 PM (10 years, 1 month ago) |
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Quote:
van der griegen said:
Quote:
Nice Ol Bud said: How'd you determine those percentages?
It's just a matter of some really easy math and knowing the density of water. Fortunately, the beautiful thing about the metric system is that we use water so much that literally every unit of measurement is based on water, so here's something easy to remember:
1cm^3 (or cc or cubic centimeter) of water is 1mL. 1mL of water weighs 1g at room temperature. Every degree Celsius is 1% of the difference between boiling water and freezing water, and a calorie is how much energy you need to raise a gram(or cc or mL) of water one degree Celsius, but that's not really relevant. And despite this, people say still SAE is easier. 
So let's say you have 5g of starches (which, as you can see from my earlier post, is actually 5g of simple sugars strung together in chains). You also have 7g of dextrose. 5g + 7g is 12g. So what percentage of the total solution is sugars?
First we need to ask ourselves, "Roughly how much will a 500mL solution weigh?" Since every mL of water weighs a gram, the answer is obviously 500g water + 12g sugars, for a total of 512g. Then just a little simple math:
12g sugars = 2.34% sugars. 512g solution
Does that help?
Quote:
Oh, it matters very much. 10mL of karo or honey is roughly 7g of dextrose/glucose, and almost all of the potato is starch. Together that brings them to 2.4% available sugars, which is ideal for your purposes. With mushrooms, any less and they don't have enough nutrients to properly grow, and any more and they see no reason to. Too much sugar result in slow, fluffy, tomentose growth which is harder to isolate and more prone to contamination as a result.
2-3% is perfect for both agar and sugar concentration. Any more or less and you stray from the ideal.
cool this is like golden info going in my agar notes
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invitro


Registered: 05/03/13
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Re: Agar Formula Help [Re: Psilicon]
#19324801 - 12/25/13 08:31 AM (10 years, 1 month ago) |
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Van Der Griegen,
if 2-3% sugar is the best growth medium, why do they say the best LC is 2% LME 2% Dextrose?
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Psilicon
Really Nice Guy


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Re: Agar Formula Help [Re: invitro]
#19324917 - 12/25/13 09:15 AM (10 years, 1 month ago) |
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I'm not sure who says that, but I also tend to make my liquid cultures on the sugary side. The more sugar you have for the fungi to use as a carbon and energy source, the more mycelial mass can be produced, right? And with an LC, the time it takes to make it is less important than the strength of the inoculant.
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Violet



Registered: 12/06/11
Posts: 4,205
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Re: Agar Formula Help [Re: Psilicon]
#19325274 - 12/25/13 11:37 AM (10 years, 1 month ago) |
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Thanks van der griegen, some good stuff!
But relevant to the use of Starch and/or sugar in media... the value of starch is that they have to make that more difficult conversion into easily usable sugars. Starch doesn't have nearly the same effects with increased density that sugars do. It's why 8% sugar is getting too hot, but 100% grain can be ripped right thru. I grow mushrooms on agar with only 1/3 of the agar-agar but with 3 times the grain nutrition - far from being "too nutritious" they still colonize like mad and fruit like crazy, although it's true they much prefer to pin away from the nutrition source. Rather than slowing down colonization due to high starch nutrition like sugar would, it instead simply needs more digestion time, which we know to be the case.
Growing on sugar long enough is known to be no good for culture's metabolic veracity, promoting and adjusting to a kind of laziness.
Using 100% grainwater gets wonderful growth all the time just like we would expect on grains, and ensures the culture is always hardy for our grassy grow media. And you don't even have to worry about nutrient richness just like you don't with grains, but do with sugar. Yet if you like one can still easily reduce grainwater.
My agar can be very very nutritious, but it's always 0% sweet (may be slightly "sweet" to mycelium due to my pre-ferment procedure) I always get prime growth, speed, sectoring, and the cultures stay just as vigorous.
I would understand adding sugars to LCs if I understood the desire to use LCs.
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Psilicon
Really Nice Guy


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Re: Agar Formula Help [Re: Violet]
#19325392 - 12/25/13 12:18 PM (10 years, 1 month ago) |
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Very good points, Violet. You may be on to something about the starch, and I think I'll end up checking that out. I know Pasty uses an impressive amount of potato in his agar compared to most PDA recipes, and he gets excellent results. An experiment is in order.
With regards to liquid cultures, I don't like them either. I have one going right now, though, since some say they're excellent for sclerotia-producing species. I've tried MS v-tek on my mexicana, and while I had excellent colonization, I got no stones at all. I'm not sure what the cause of it was, but the MS WBS jar I inoculated at the about the same time is colonizing very slowly while making a hell of a lot of truffles. I'm looking for a quick, low-risk inoculant, and agar-->LC-->grain could fit the bill.
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Stromrider
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Re: Agar Formula Help [Re: Psilicon]
#19325499 - 12/25/13 01:02 PM (10 years, 1 month ago) |
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I been using dog food agar that Amanita Virosa makes and sells at his Cultivation supply shop. It's been working really well for me. It's got a small amount of potato flakes and nutritional yeast along with the crushed dog food.
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36fuckin5
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It all depends on the species you want to grow. Some species don't like DFA. Some like higher or lower levels of sugars/starches.
Almost everything will grow on standard MEA or PDA mix with a 2% sugar content. FastFred's Media Cookbook has all the info you need.
Start looking in more scientific places for info about agar than here. There are many many different formulas, and all are dependent on what you want to grow.
If you don't want to go read all the little nuances, then save yourself the time and use a standard MEA or PDA. They're cheap, easy and common. There's not much use in reading more about it here, you've pretty much gotten all the info on the forums in tbis thread. There's plenty of use in reading some scientific articles on agar and different formulations.
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invitro


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Re: Agar Formula Help [Re: Violet]
#19329407 - 12/26/13 03:49 PM (10 years, 1 month ago) |
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Hi Violet,
I have soak water from boiled rice and soak water from grass seed, I'm pcing each in a syringe for injection into a lc. Some lcs have honey water which is depleted of nutrients by now I'm sure. Others have 2% each of LME/DEX which are fresher. Do you have any recommended dosages?
Thanks for the link to the rice prep and grain water use for agar. It seems like good info.
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