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Offlinehamloaf
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Any Reason Why Agar,
    #14051637 - 03/01/11 10:19 PM (12 years, 10 months ago)

don't solidify during the PC cycle? 

Just PC'ed quite a few jars of agar MEA+ for 40 minutes at 15-20 psi.  Upon opening the PC after letting the PC cool for 12 hours, not one jar of agar had solidified after cooling from the sterilization process.  Any idea what may have caused this?


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Re: Any Reason Why Agar, [Re: hamloaf]
    #14051661 - 03/01/11 10:25 PM (12 years, 10 months ago)

did you use a diff type then usual? or maybe not enough?

idk i've never had that problem ;/


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Re: Any Reason Why Agar, [Re: biologys]
    #14051691 - 03/01/11 10:30 PM (12 years, 10 months ago)

Hey biologys, thanks for the reply.  Semi perplexing agar issue, isn't it.

Nosir.  Used my normal agar regimen and PC duration (10 grams LME, 9 grams agar, .5 of a gram peptone, PC'ed for 40 minutes at 15-20 psi's).  One point of interest, about 75% of the agar jars sterilized were prepared 24 hours ago and had been sitting on the counter at room temps before they were sterilized.  25% of the jars just PC'ed were prepared freshly.  All jars didn't solidify.


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Re: Any Reason Why Agar, [Re: hamloaf]
    #14051754 - 03/01/11 10:42 PM (12 years, 10 months ago)

hmm, thats really weird

I normally mix all ingredients, boil pour in jars and let sit for 24 hours before PC'n myself..and no problems so far, did it solitify during the 24 hour sit before PC'n?


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Re: Any Reason Why Agar, [Re: biologys]
    #14051774 - 03/01/11 10:45 PM (12 years, 10 months ago)

same jar lids?

foil over jar lids?

only thing i could think of is too much water, whether or not it was introduced prior to cooking couldnt say


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Re: Any Reason Why Agar, [Re: biologys]
    #14051781 - 03/01/11 10:45 PM (12 years, 10 months ago)

Quote:

did it solitify during the 24 hour sit before PC'n?



No.


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Re: Any Reason Why Agar, [Re: hamloaf]
    #14051793 - 03/01/11 10:47 PM (12 years, 10 months ago)

idk bro, wish i could help out, but ill def be watching to see if someone with more experience gives answer to this..


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Re: Any Reason Why Agar, [Re: hamloaf]
    #14051802 - 03/01/11 10:49 PM (12 years, 10 months ago)

Quote:

hamloaf said:
Hey biologys, thanks for the reply.  Semi perplexing agar issue, isn't it.

Nosir.  Used my normal agar regimen and PC duration (10 grams LME, 9 grams agar, .5 of a gram peptone, PC'ed for 40 minutes at 15-20 psi's).  One point of interest, about 75% of the agar jars sterilized were prepared 24 hours ago and had been sitting on the counter at room temps before they were sterilized.  25% of the jars just PC'ed were prepared freshly.  All jars didn't solidify.




Bro, you add a few extra constraints at the end there that threw me.

Weird.

I have just done agar by the numbers (yours actually 9/10 ;0) and have
not had to reheat any.  I have gotten the impression that agar can be
heated and cooled more than once.  I do not know.  Sorry!

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Re: Any Reason Why Agar, [Re: k00laid]
    #14051807 - 03/01/11 10:49 PM (12 years, 10 months ago)

Quote:

k00laid said:
same jar lids?

foil over jar lids?

only thing i could think of is too much water, whether or not it was introduced prior to cooking couldnt say



Hey k00l-aid, yes.  Used my usual jar lids and foil was placed thoroughly over jar lids all the way over to where the lid bands meet the glass jar.  I'm tellin' you, didn't change a thing about the way I normally prepare agar.


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Re: Any Reason Why Agar, [Re: biologys]
    #14051813 - 03/01/11 10:50 PM (12 years, 10 months ago)

well then.... :popcorn:


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Re: Any Reason Why Agar, [Re: biologys]
    #14051850 - 03/01/11 10:54 PM (12 years, 10 months ago)

u need to test the pH.  pH needs between 5-6 to solidfy.  old agar will have trouble solidifying too.


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Re: Any Reason Why Agar, [Re: k00laid]
    #14051856 - 03/01/11 10:55 PM (12 years, 10 months ago)

If 25% of the jars just PC'ed were prepared freshly, then the delay for
the other 75% was not an issue.

Curiouser and curiouser...

JD


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Re: Any Reason Why Agar, [Re: dmonkey1]
    #14051869 - 03/01/11 10:57 PM (12 years, 10 months ago)

Quote:

dmonkey1 said:
pH needs between 5-6 to solidfy.




im skeptical but so far this is the only answer xDD


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Re: Any Reason Why Agar, [Re: dmonkey1]
    #14051878 - 03/01/11 10:58 PM (12 years, 10 months ago)

none solidified he said?  The mix could be on the basic side instead of acidic.


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Re: Any Reason Why Agar, [Re: k00laid]
    #14051897 - 03/01/11 10:59 PM (12 years, 10 months ago)

Quote:

k00laid said:
Quote:

dmonkey1 said:
pH needs between 5-6 to solidfy.




im skeptical but so far this is the only answer xDD





why do you think jokefox's agar broth doesn't solidify after treatment, yet it will solidify before treatment?


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Re: Any Reason Why Agar, [Re: dmonkey1]
    #14051908 - 03/01/11 11:01 PM (12 years, 10 months ago)

I would do another small test run to see if you accidentally made a mistake with measurements. Likely not the case but that will tell you if it is product or operator error.:shrug:


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Re: Any Reason Why Agar, [Re: dmonkey1]
    #14052012 - 03/01/11 11:15 PM (12 years, 10 months ago)

Quote:

need to test the pH.  pH needs between 5-6 to solidfy.  old agar will have trouble solidifying too.




Hey dmonkey1.  Sounds like you maybe on to a couple of solutions.  How old is too old for agar, or what's agar's shelf life? 

The agar being old makes the most sense.  Had this particular bottle of agar for about 3-4 months.  Made some agar jars last week from it that were good using the same recipe always used so the agar could'v just reached it's age limit in the last 7 days.  About the PH, like I said, never had this problem before but I'm open to the idea that the PH maybe the issue although it doesn't make sense being that I haven't changed a thing about the way I make agar.  How and when is agar tested for PH?


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Re: Any Reason Why Agar, [Re: hamloaf]
    #14052042 - 03/01/11 11:21 PM (12 years, 10 months ago)

old agar = many years old.  I have 5 year old agar that I use for LC hahaha


I'd add an acid to the agar.  pH paper is ideal method.  or purple cabbage juice.  I'd add 1mL portions of whatever you use (vinegar is weak acid, should do you fine) until it solidifies if u wanna play guess to get it in the slightly acidic range (5-6pH)


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Re: Any Reason Why Agar, [Re: dmonkey1]
    #14053123 - 03/02/11 07:33 AM (12 years, 10 months ago)

:strokebeard:


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Re: Any Reason Why Agar, [Re: hamloaf]
    #14053420 - 03/02/11 09:08 AM (12 years, 10 months ago)

Quote:

hamloaf said:
don't solidify during the PC cycle? 

Just PC'ed quite a few jars of agar MEA+ for 40 minutes at 15-20 psi.  Upon opening the PC after letting the PC cool for 12 hours, not one jar of agar had solidified after cooling from the sterilization process.  Any idea what may have caused this?




This is not surprising.  My plates don't solidify until they've been at room temperature for a few hours.  Even cool plates are still semi-liquid until that amount of time has passed.
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Re: Any Reason Why Agar, [Re: RogerRabbit]
    #14053529 - 03/02/11 09:32 AM (12 years, 10 months ago)

Hey RogerRabit, thanks.  I didn't throw away the agar yesterday rather I placed the jars in a large storage tote to see if anything would change.  Today every single jar's agar is still liquid.

Thanks for everyones help and input on the matter of my agar fuck-ups.  I'mma try it again in a few hours once the grains are done boiling.  I'll be letting you all know what happens to the next round of agar prepared.


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Re: Any Reason Why Agar, [Re: hamloaf]
    #14054303 - 03/02/11 12:15 PM (12 years, 10 months ago)

i'd buy the good stuff.  commercial agar is the bee's knees.  Mine hardens in like 30-40 minutes so I leave it under the running hood and do work when its ready :smile: safes me the headache of dealing with blank dishes


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Re: Any Reason Why Agar, [Re: dmonkey1]
    #14054471 - 03/02/11 12:55 PM (12 years, 10 months ago)

Ok, an interesting discovery was made.  Broke into the tub to clean and reload the jars then begin again to discover the jars of agar that had been freshly prepared all had solidified.  Every single one. 

I know that they were the jars freshly prepared because they were landmarked in case I had an issue with leaving the jars of agar on the counter mixed into water 24 hours before I PC'ed it.  I should have my head examined, for that was the only thing about my agar regimen that I changed and kind of had a pre-emptive bad feeling about.  While not one of the jars that sat on the counter before sterilization for 24 hours did ever solidify.

dmonkey1, I use half-pint wide mouth jars for agar plates/dishes.  The agar is pre-mixed and poured into the jars and PC'd at once.  Allotting a 12-24 hour cool down of the PC before removing the ready to be inoculated agar dishes.  Although I have a flowhood, doing agar this way enables you safely inoculate agar in open air with a syringe solution.  :thumbup:


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Re: Any Reason Why Agar, [Re: hamloaf]
    #14054569 - 03/02/11 01:18 PM (12 years, 10 months ago)

Quote:

hamloaf said:
Ok, an interesting discovery was made.  Broke into the tub to clean and reload the jars then begin again to discover the jars of agar that had been freshly prepared all had solidified.  Every single one. 

I know that they were the jars freshly prepared because they were landmarked in case I had an issue with leaving the jars of agar on the counter mixed into water 24 hours before I PC'ed it.  I should have my head examined, for that was the only thing about my agar regimen that I changed and kind of had a pre-emptive bad feeling about.  While not one of the jars that sat on the counter before sterilization for 24 hours did ever solidify.

dmonkey1, I use half-pint wide mouth jars for agar plates/dishes.  The agar is pre-mixed and poured into the jars and PC'd at once.  Allotting a 12-24 hour cool down of the PC before removing the ready to be inoculated agar dishes.  Although I have a flowhood, doing agar this way enables you safely inoculate agar in open air with a syringe solution.  :thumbup:




I'd imagine cloning to be a nightmare with those jars :laugh:


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Re: Any Reason Why Agar, [Re: dmonkey1]
    #14054741 - 03/02/11 01:46 PM (12 years, 10 months ago)

Quote:

I'd imagine cloning to be a nightmare with those jars



This is going to depend largely on the method you use for cloning.  Although, generally yes.  Cloning or any agar work is easier in petri dishes.  If you have a flowhood then these jars are a cinch to work with when cloning no matter what method of cloning you use.


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Re: Any Reason Why Agar, [Re: hamloaf]
    #14054766 - 03/02/11 01:51 PM (12 years, 10 months ago)

Quote:

hamloaf said:
Quote:

I'd imagine cloning to be a nightmare with those jars



This is going to depend largely on the method you use for cloning.  Although, generally yes.  Cloning or any agar work is easier in petri dishes.  If you have a flowhood then these jars are a cinch to work with when cloning no matter what method of cloning you use.




IME,  cloning will have contams associated.  Isn't more volume between media and the lid a very very bad thing?  I can only imagine more volume = more bad spores


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Re: Any Reason Why Agar, [Re: hamloaf]
    #14058869 - 03/03/11 09:11 AM (12 years, 10 months ago)

Freshly prepared some more agar and directly into the PC it went for 40 minutes at 15-20psi's.  Let them cool in the PC for 14 hours before opening the PC.  All jars that were recently PC'ed have solidified.  I'mma have to credit this mistake to preparing the agar 24 hours in advance and leaving it on the counter at room temps before PC'ing, being as that's the only thing I changed about my normal agar regimen.


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Re: Any Reason Why Agar, [Re: hamloaf]
    #14058922 - 03/03/11 09:26 AM (12 years, 10 months ago)

thats definately weird, every jar that i've prepped since i've started with agar has been left to sit out for a day atleast before PC'n...:shrug: who knows glad your prob is fixed :smile:


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Re: Any Reason Why Agar, [Re: biologys]
    #14058970 - 03/03/11 09:42 AM (12 years, 10 months ago)

Most people who use non-lab-grade agar boil it in water before adding the malt or other nutrient solution, followed by sterilization.  If you're using grocery store agar flakes, this could have been the problem.

I have so much to do that there's no time for all that.  I still use fungi perfecti agar mix simply because it takes five minutes to go from powder to the pressure cooker, and then I pour when it's cool enough to handle, usually 2 hours after the pressure cooker returns to zero pressure.  It's never failed me yet.
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Re: Any Reason Why Agar, [Re: RogerRabbit]
    #14059005 - 03/03/11 09:53 AM (12 years, 10 months ago)

Quote:

RogerRabbit said:
Most people who use non-lab-grade agar boil it in water before adding the malt or other nutrient solution, followed by sterilization.




That's what I was lead to believe I should do, having read that the agar needs to reach a boil in the water to take on its desirable qualities.  I never tested the necessity of the boil, but it always works :grin:

I put the water in a pot, add agar powder and extras while it's cold, heat and whisk it until it boils (don't let it boil over, whatever you do), strain, and PC.


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Re: Any Reason Why Agar, [Re: andymc]
    #14059390 - 03/03/11 11:18 AM (12 years, 10 months ago)

I also use FP MEA


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Re: Any Reason Why Agar, [Re: RogerRabbit]
    #14059572 - 03/03/11 11:53 AM (12 years, 10 months ago)

RR Just out of curiosity how much agar pre-mix do you go through in
any given week/month? How many plates do you find yourself pouring
on a week/month basis to keep your shiitake operation going?


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Re: Any Reason Why Agar, [Re: RogerRabbit]
    #14074360 - 03/06/11 06:16 AM (12 years, 10 months ago)

Quote:

Most people who use non-lab-grade agar boil it in water before adding the malt or other nutrient solution, followed by sterilization.  If you're using grocery store agar flakes, this could have been the problem.



NOW brand, grocery store agar was used.  This agar is the consistency of powder, not flakes.  Didn't boil the water before adding the malt or other nutrients before sterilization as I never do.  Use luke warm water to mix the ingredients into and make the agar solution before sterilization though. 

Not into the production range or level you are.  Hense the mixing of my own agar, as well as, the use of half-pint wide mouthed jars instead of petri dishes for culturing mycelium.  When that day comes, premixed agar from a sponsor'll be kept in the cultivation supply area, as well as, petri dishes.  lol

Thanks, bio.


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