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OfflineKingCloyster
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Registered: 05/27/13
Posts: 597
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Problems with spore syringe not germinating on LME agar.
    #19061045 - 10/30/13 11:13 PM (10 years, 2 months ago)

I use 5gs agar and like 2.5 to 4 gs of LME per 250ml of water. 17-18 PSI for 40 minutes. Pour, stacked, blah, no condensation, blah, blah. Then 2 days later I knock up 4 plates of agar, with 3 different syringes and parafilm them under a flowhood... Every single time, I get a distinct bacterial contamination instead of mycelium.  Its always shiny milky colored or shiny pink. Usually right around where i dropped the spore solution.

I have done this 4 times now for a total of 15-20 plates, and every single time i dont get germination. just slimy bacteria.... Is my agar too wet? are all of my syringes contaminated? shouldnt the spore still germinate?


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OfflinePussyFart
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Re: Problems with spore syringe not germinating on LME agar. [Re: KingCloyster]
    #19061085 - 10/30/13 11:20 PM (10 years, 2 months ago)

Quote:

KingCloyster said:
are all of my syringes contaminated? shouldnt the spore still germinate?



Sounds like it.

Yes, the spores should still germinate.

Kinda like this.

       


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OfflineKingCloyster
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Re: Problems with spore syringe not germinating on LME agar. [Re: PussyFart]
    #19062702 - 10/31/13 10:47 AM (10 years, 2 months ago)

Fuck so I need to order new spores huh?, I figured u could clean them up


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OfflineKingCloyster
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Registered: 05/27/13
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Re: Problems with spore syringe not germinating on LME agar. [Re: KingCloyster]
    #19062773 - 10/31/13 11:02 AM (10 years, 2 months ago)

Why would the syringes not be germinating? Temperature swings possibly?


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OfflinePussyFart
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Re: Problems with spore syringe not germinating on LME agar. [Re: KingCloyster]
    #19062794 - 10/31/13 11:05 AM (10 years, 2 months ago)

You can....that is one of the many great things about agar...you can transfer away from contams.

I did it with those very dishes....just wait till something germinates and grows enough to where you can cut a clean wedge.

That is if the bacteria does not keep growing....How long does it take for the bacteria to show up, and how long after that did you keep them?

Because spores can take up to 2+ weeks to germinate in some cases....


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Invisiblepoopy mcpooperson
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Re: Problems with spore syringe not germinating on LME agar. [Re: PussyFart]
    #19063081 - 10/31/13 12:15 PM (10 years, 2 months ago)

i have been having the same problem and i just put print spores on some agar this morning. i have a thread that i used all spores from various MS syringes and they almost all were contaminated. i used way too much solution when i went to agar the first time too. you can tell from my plates that the spore solution went all over the petri dishes.

i was told that syringes are better just to inoculate jars and then go grain to agar from those fully colonized jars. i am going to do the grain dunk TEK to agar because from what ive seen it takes off faster and has better growth. good luck. take a look at all my photos of contaminated agar from MS syringes if youd like. there is some good info from members guiding me in the right direction. ill also attach the link to grain dunk to agar.

my agar log:
http://www.shroomery.org/forums/showflat.php/Number/19013539#19013539

franks grain dunk TEK:
http://www.shroomery.org/forums/showflat.php/Number/18781503#18781503


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love and laughter  :laugh:


Edited by poopy mcpooperson (10/31/13 12:55 PM)


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OfflineKingCloyster
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Re: Problems with spore syringe not germinating on LME agar. [Re: poopy mcpooperson]
    #19063463 - 10/31/13 01:30 PM (10 years, 2 months ago)

It takes 2 days for the pink some 3 to 5 days for the rest.. I don't keep them much longer after that. Maybe a week...

It looks as if some did germinate but were covered in a liquid still. Perhaps the syringe solution?  It looks very similar to ur Colorado ms syringe in your Agar log:
I just don't see fuzz. It all looks to be slimy


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Offlinebotanicalbishop
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Re: Problems with spore syringe not germinating on LME agar. [Re: KingCloyster]
    #19063980 - 10/31/13 03:14 PM (10 years, 2 months ago)

Did you make your syringes or buy them for a trusted vendor? Double check that your flowhood is working properly whether its the filter or the carbon scrubber. If you have an SAB laying around try that and see if you get the same results, if not try moving your flow hood into a different room. If all else fails just buy another syringe there not expensive, unlike the precious time you waste trying to battle contams. Good luck


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"Prohibition will work great injury to the cause of temperance. It is a species of intemperance within itself, for it goes beyond the bounds of reason in that it attempts to control a man's appetite by legislation, and makes a crime out of things that are not crimes. A Prohibition law strikes a blow at the very principles upon which our government was founded."

Abraham Lincoln


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OfflineKingCloyster
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Re: Problems with spore syringe not germinating on LME agar. [Re: botanicalbishop]
    #19091528 - 11/05/13 06:13 PM (10 years, 2 months ago)

Is it best to have the flowhood against the back wall in a small room?


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OfflineKingCloyster
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Registered: 05/27/13
Posts: 597
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Re: Problems with spore syringe not germinating on LME agar. [Re: KingCloyster]
    #19091916 - 11/05/13 07:18 PM (10 years, 2 months ago)

Oh and I bought them from mushroom prints.com.  so far, Germination ratio is less than 20%


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OfflineKingCloyster
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Registered: 05/27/13
Posts: 597
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Re: Problems with spore syringe not germinating on LME agar. [Re: KingCloyster]
    #19095985 - 11/06/13 03:30 PM (10 years, 2 months ago)

Bump


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Offline1down5up
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Re: Problems with spore syringe not germinating on LME agar. [Re: KingCloyster]
    #19354510 - 01/01/14 12:13 PM (10 years, 29 days ago)

Do you still have the syringes?  You can test them on some other medium to see if it's them.  At least that way you can rule out one option....


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