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OfflineRotnpins
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Let's talk about Water Agar!! ๐Ÿ’ง * 31
    #27900393 - 08/12/22 10:03 AM (1 year, 5 months ago)

When I first started working with agar, I had an abundance of plates that I thought were too contaminated to clean up.. I thought about using antibiotics, but i had read that it could also hinder the mycelium growth and I didn't want to get stuck using it as a crutch.. So I decided to try making some water agar..

What is "water agar"?
Water agar is exactly what it sounds like, water and agar without any added nutrients.. the mycelium grows out very quickly (compared to the contams) looking for food.. this gives it a chance to out grow any contamination that may be riding along.

How to make water agar:
The recipe I use is 7g Agar powder per 500ml H2O

I leave the plates clear and add a different food coloring to each of my nutritional plates so I can easily distinguish between the type of plates.. (for example: H2O Agar=clear, MEA= blue, YMEA= yellow, etc,.)



You would prepare the water agar the same way as your nutritional agar plates.. the method I use is this:

1: add 100ml cold/room temp H20 to media bottle.
2: measure out remaining water (400ml for a 500ml batch) and heat water using preferred method. I personally bring it to a light boil on the stove.
3: while the water is heating, measure out your agar powder (7g per 500ml) and add it to your media bottles. Then give the bottle a little bit of a shake/stir.
4: once it is hot, add the remaining water to the media bottle. Put the lid on and shake it up.
5: prepare your PC with enough water for a 20 minute cycle.. put the water agar media bottle in the pc. If you have any nutritional agar, instruments that need sterilizing, no pour plates, etc,. you can throw them in the PC at the same time.
6: vent steam from PC for 10-15 minutes, add your weight, bring PC up to 15-17psi, reduce heat to low,, set a timer โฒ and let cook for 20 minutes.
7: when PC cycle is done, turn the heat off and wait for pressure to drop to 0 on its own.
8: take your bottle(s) to your work space and wait for them to cool to pouring temp (I usually pour between 140-120 degrees)

How to use water agar:
Once your plates have cooled, it's time to do some transfers..

I make my transfers the same way I would when transferring to a nutrient plate.. I find an area that seems to have the most organized growth and I take a small transfer to the new plate.. smaller transfer wedges may be a little more difficult to work with at first, but they are less likely to carry contamination to the new plate. So try practicing taking smaller transfers each time until you're able to easily transfer a piece that is about the size of a grain of rice.

Wait for the mycelium to grow out.. it will be very thin and whispy, and it will grow out toward the edges much quicker than any contamination that tags along.

When the mycelium has grown out enough that I feel confident it has outgrown the contamination (its a good idea to let it at least half way to the edge, or all the way to the edge for more piece of mind), I take a few transfers from the water agar plate and move them to nutritional plates.

Next step is easy: watch your new plate as the mycelium grows out clean!

Edit:
Here's a video I found that discusses a couple applications for water agar that I didn't mention



Pictures added 8/28:

Here are some pictures to show you guys the growth on a few plates.. they aren't the best pictures,  but you should be able to see the leading edge to get an idea of the speed of growth.

Plate stared 8/10 (18 days ago):



8/11 (17 days ago):



8/12 (16 days ago):




Agar to grain comparison
Water Agar vs Nutrient Agar

10/28:
I'm on a mission to start using up some of my ridiculously large plate collection.. So, I decided to do a side-by-side, plate to grain, comparison using 2 fully grown out Phobos plates.. One plate is water agar, the other is MYA. I decided to do 3 jars from each plate.

Quick picture before getting to work:


Here are the plates I'll be using


Plate 1 (MYA- cut into 3 wedges, leaving the center and outer ring behind:


Used plate 1 to inoculate 3 jars of oats:


Here is plate 2 (WA- cut in a similar pattern to plate 1)


Used plate 2 to inoculate the other 3 oat jars:


I plan to post updates comparing any differences in the length of time it takes for the mycelium to jump off, how long it takes the jars to colonize, etc...

11/3

The WA plates are finally starting to jump off to the grain a little bit.. they are definitely running a few days behind the jars that were inoculated using nutrient agar.. like I said in a previous post, this could be due to the age of the plates.. it is a possibility that the mycelium was in a dormant state, due to the fact that they're around 3 months old.. I still plan to run another test with fresh plates to see if the results differ from this batch.

Enigma (WA)


Phobos (WA)


Phobos (MYA)



Edited by Rotnpins (11/03/22 10:56 AM)


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OfflineLand TroutM
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Re: Let's talk about Water Agar!! ๐Ÿ’ง [Re: Rotnpins] * 1
    #27900403 - 08/12/22 10:13 AM (1 year, 5 months ago)

Cool man, thanks for sharing.๐Ÿ’ฆ๐Ÿ’ฆ


Edited by Land Trout (08/12/22 10:13 AM)


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InvisibleBigwormS
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Re: Let's talk about Water Agar!! ๐Ÿ’ง [Re: Land Trout] * 1
    #27900421 - 08/12/22 10:21 AM (1 year, 5 months ago)

Cool write up.


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OfflineSmellyhobbit
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Re: Let's talk about Water Agar!! ๐Ÿ’ง [Re: Rotnpins] * 1
    #27900437 - 08/12/22 10:29 AM (1 year, 5 months ago)

Good write up for something not in widespread use. People will find this useful. Nice work


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Offlinesan pedro guy
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Re: Let's talk about Water Agar!! ๐Ÿ’ง [Re: Smellyhobbit] * 1
    #27900523 - 08/12/22 11:36 AM (1 year, 5 months ago)

nice!

have you tried transferring some bacteria and myc on the same plate?

Im also curious if the water agar would work better or worse for sandwhich agar


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OfflineRotnpins
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Re: Let's talk about Water Agar!! ๐Ÿ’ง [Re: san pedro guy]
    #27900540 - 08/12/22 11:52 AM (1 year, 5 months ago)

Quote:

san pedro guy said:
nice!

have you tried transferring some bacteria and myc on the same plate?

Im also curious if the water agar would work better or worse for sandwhich agar





Yes.. I've transferred mycelium that's grown through bacteria and its enmeshed. It works really well for bacterial mycelium..

I actually just started a bunch of new water agar transfers yesterday so I can update with some pictures showing the process and how the nutritional plates look before and after cleaning up with water agar.. so hopefully I'll be updating in a few days with pictures showing the process


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Offlinewild_aussie
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Re: Let's talk about Water Agar!! ๐Ÿ’ง [Re: Rotnpins] * 1
    #27900587 - 08/12/22 12:37 PM (1 year, 5 months ago)

Great write up just really getting into agar myself and have had issues with slow growth where bacteria had a chance to take hold can't wait to try this.
:myman:


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OfflineRotnpins
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Re: Let's talk about Water Agar!! ๐Ÿ’ง [Re: wild_aussie] * 1
    #27900598 - 08/12/22 12:44 PM (1 year, 5 months ago)

Quote:

wild_aussie said:
Great write up just really getting into agar myself and have had issues with slow growth where bacteria had a chance to take hold can't wait to try this.
:myman:




For sure! I've only done a couple test runs so far, but the results have been really good... I have a feeling that this will be a game changer.. I'm keeping water agar plates on standby so I've got a reliable tool ready whenever I've got a contaminated plate that needs some extra TLC.

If anyone gives it a try, I'd really appreciate it if you shared your experiences and photos in this thread so everyone can see the results from multiple people :mushroom2:


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InvisibleHILLBILLY OUTLAWS
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Re: Let's talk about Water Agar!! ๐Ÿ’ง [Re: Rotnpins] * 1
    #27900624 - 08/12/22 01:03 PM (1 year, 5 months ago)

Love the write up. Very useful tek for somebody like me thatโ€™s just getting into agar and struggling. Lol gonna do some more plates this weekend. Iโ€™ll give water agar a shot as well.

:headbang3:


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Re: Let's talk about Water Agar!! ๐Ÿ’ง [Re: Rotnpins] * 1
    #27900626 - 08/12/22 01:05 PM (1 year, 5 months ago)

I meant more like transfer myc on one part of the plate then transfer some bacteria to the other side (or a separate plate) to see how each responds to the nutritional deprivation

either way iโ€™ll be following; good shit rotn :thumbup:


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OfflineRotnpins
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Re: Let's talk about Water Agar!! ๐Ÿ’ง [Re: san pedro guy]
    #27900645 - 08/12/22 01:17 PM (1 year, 5 months ago)

Quote:

san pedro guy said:
I meant more like transfer myc on one part of the plate then transfer some bacteria to the other side (or a separate plate) to see how each responds to the nutritional deprivation

either way iโ€™ll be following; good shit rotn :thumbup:





That's a good idea ๐Ÿ’ก I don't have any plates with bacterial colonies that haven't been overtaken by mycelium at the moment, but I'm sure I'll come across some soon and give it a try and post the results


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Re: Let's talk about Water Agar!! ๐Ÿ’ง [Re: Rotnpins] * 1
    #27900720 - 08/12/22 01:54 PM (1 year, 5 months ago)

Quote:

That's a good idea ๐Ÿ’ก I don't have any plates with bacterial colonies that haven't been overtaken by mycelium at the moment, but I'm sure I'll come across some soon and give it a try and post the results




Shouldn't be too hard to find bacteria.  Grab your kitchen sponge, open one of your plates, touch the corner of the sponge to the agar, and seal her back up.  Or, I can send you a complimentary plate in the name of science!  Lol.

On a more serious note, I see huge potential for this as being a 2 transfer until clean, opposed to a 3 transfer until clean.  That will literally save days, weeks, and for some of you old timers, years in transfer/wait time.

I'll definitely be following this post in the future.  Nice job, Rotnpins!


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OfflineRotnpins
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Re: Let's talk about Water Agar!! ๐Ÿ’ง [Re: zilker]
    #27901235 - 08/12/22 07:23 PM (1 year, 5 months ago)

Quote:

zilker said:
Quote:

That's a good idea ๐Ÿ’ก I don't have any plates with bacterial colonies that haven't been overtaken by mycelium at the moment, but I'm sure I'll come across some soon and give it a try and post the results




Shouldn't be too hard to find bacteria.  Grab your kitchen sponge, open one of your plates, touch the corner of the sponge to the agar, and seal her back up.  Or, I can send you a complimentary plate in the name of science!  Lol.

On a more serious note, I see huge potential for this as being a 2 transfer until clean, opposed to a 3 transfer until clean.  That will literally save days, weeks, and for some of you old timers, years in transfer/wait time.

I'll definitely be following this post in the future.  Nice job, Rotnpins!





:laugh2: true.. but like you said, it's not too difficult to find bacteria. I'm sure I'll have some on at least one of my germ plates soon :lol:


Exactly ๐Ÿ’ฏ  I completely agree...my goal, over time, is to consistently reduce the number of transfers needed, even when starting with heavy contamination.


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Offlinerumfor69
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Re: Let's talk about Water Agar!! ๐Ÿ’ง [Re: Rotnpins] * 1
    #27901299 - 08/12/22 08:20 PM (1 year, 5 months ago)

Nice write up I could have definitely used this many times in the past. Is the reason to use 7g agar instead of 10g so that it's a little softer?


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OfflineBagOfDicks
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Re: Let's talk about Water Agar!! ๐Ÿ’ง [Re: zilker] * 1
    #27901331 - 08/12/22 08:40 PM (1 year, 5 months ago)

I'll definitely be giving this a go. ๐Ÿ’ก


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Re: Let's talk about Water Agar!! ๐Ÿ’ง [Re: Rotnpins] * 1
    #27901386 - 08/12/22 09:22 PM (1 year, 5 months ago)

I have one RustyWhyte syringe that's contaminated with bacteria which I've been holding on to in hopes of cleaning it somehow. I might have to try this method.


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InvisibleBaba Yaga
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Re: Let's talk about Water Agar!! ๐Ÿ’ง [Re: WellDuYaPunk] * 1
    #27901435 - 08/12/22 10:18 PM (1 year, 5 months ago)

Cool stuff, will try this with clone tissue, will be interesting to see if that will
make the washing wish H2O2 or bleach obsolete.

:thumbup::thumbup::thumbup::thumbup::thumbup:


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OfflineGrind365
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Re: Let's talk about Water Agar!! ๐Ÿ’ง [Re: Baba Yaga]
    #27901468 - 08/12/22 11:08 PM (1 year, 5 months ago)

So would you use this all the time or only when you need to clean up a culture?


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OfflineRotnpins
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Re: Let's talk about Water Agar!! ๐Ÿ’ง [Re: Grind365]
    #27901618 - 08/13/22 05:48 AM (1 year, 5 months ago)

Quote:

Baba Yaga said:
Cool stuff, will try this with clone tissue, will be interesting to see if that will
make the washing wish H2O2 or bleach obsolete.

:thumbup::thumbup::thumbup::thumbup::thumbup:




Awesome! You should post your results here so we can see how it goes


Quote:

Grind365 said:
So would you use this all the time or only when you need to clean up a culture?




I would just do this for clean up..

If you're wanting a bit quicker growth on your nutritional plates, you can always try out a low nutrient level.


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OfflineAwestruck
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Re: Let's talk about Water Agar!! ๐Ÿ’ง [Re: Rotnpins] * 1
    #27902234 - 08/13/22 05:33 PM (1 year, 5 months ago)

That's a really cool, novel idea. Sounds good. Thanks for sharing the tek!


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