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OfflineKaydren
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Liquid Culture help
    #4207600 - 05/22/05 08:36 PM (11 years, 7 months ago)

Hey, I tried Agar's badass Easy Liquid Culture tek
http://www.shroomery.org/forums/showflat.php/Cat/0/Number/4141612/an/0/page/2

Now, I injected spores and such, but theyve been incubating for about a week and I still see little black spores sitting around in the jar...  Its still clear water so I dont think I caramelized the Karo... but is there a reason theres not even a little bit? 
I tried The Super Easy Karo Tek!
http://www.shroomery.org/forums/showflat.php/Cat/0/Number/2321056/page//fpart/all/vc/1 

It grew, but it only grew a few strands and its now been sitting around for awhile, and still no more growth :frown:  Am I doing something wrong?  and should I try a b2b (bottle to bottle, Im so clever! :P ) transfer?  Suck up a strand or two of the other mycelium and inject it into the baby bottle? or is that most likely to a) not work becuase whatever is wrong with the other LC will be wrong with the new one or b) contam ?  Thanks guys, Im just getting anxious because I dont seem to have any spore solution left and I have WBS that neeeeeds to be knocked up! :frown:  Thanks!

p.s.  I followed both teks to the letter :frown:


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OfflineVALIS
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Re: Liquid Culture help [Re: Kaydren]
    #4207922 - 05/22/05 10:13 PM (11 years, 7 months ago)


It's possible that you either got the amount of karo wrong, or you didn't stir the karo well enough into the water.

How long has it been sitting w/ no growth?

How many cc's spore solution did you inject?

Have you aerated the jar? Did you shake it vigorously to oxygenate the water?

Are you certain that it's consistently incubating at the proper temps?


There's ways you can "save" the bottle, but first you need to know what the likely problem may be.


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Nature is the Technology of the Divine.


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Invisibleagar
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Re: Liquid Culture help [Re: Kaydren]
    #4207949 - 05/22/05 10:21 PM (11 years, 7 months ago)

You should see growth inside 2 weeks? You sure the syringe was good?


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OfflineLethalX5
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Re: Liquid Culture help [Re: agar]
    #4208166 - 05/22/05 11:22 PM (11 years, 7 months ago)

Quote:

agar said:
You sure the syringe was good?




i was thinking that also


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OfflineVALIS
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Re: Liquid Culture help [Re: LethalX5]
    #4208390 - 05/23/05 12:19 AM (11 years, 7 months ago)

Quote:

LethalX5 said:
Quote:

agar said:
You sure the syringe was good?




i was thinking that also





I've had a perfectly good syringe not produce results in a couple LC jars before; while other LC's from the same syringe went just fine. Syringe was shaken well each time and the same tek was followed.

I've also had a bum syringe that didn't produce jack shit, period.


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OfflineKaydren
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Re: Liquid Culture help [Re: Kaydren]
    #4210073 - 05/23/05 03:01 PM (11 years, 7 months ago)

Its been sitting without growth for a little while, Id say since about last monday or tuesday, 2 ccs spore solution per bottle and like Valis said    its a perfectly good syringe asfar as I can see, because the same syringe produced myc growth in the other LC and its from Hawks Eye, but Im not sure, and they werent at consistent temps :frown: which couldve messed it up a bit :frown: now theyre at a consistant temp tho


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OfflineVALIS
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Re: Liquid Culture help [Re: Kaydren]
    #4210306 - 05/23/05 03:59 PM (11 years, 7 months ago)

Ok - here's what to do ( assuming you want to put in the effort of trying to revive it, rather than just making a new one ):

You've gotten the temps stabilized which is good ( though that shouldn't have caused too big an issue, unless the temp was fluctuating pretty wildly ).

Next thing, get an empty sterilized syringe. Using a lighter, place the needle into the flame and aspirate fully with air. Wait for the needle to cool momementarily, swab it with some alcohol then quickly inject this into your LC, bubbling and aerating the culture water. Take out the syringe, then shake the hell out of the bottle ( you have a marble in there right? ).

Now you know there's no lack of oxygen for the proper growth of the myc. Put back into incubator for a couple days ( about three ), if you still don't see growth, then try the following:

Get a sterilized needle ready. Boil a very small amount of distilled water ( about 100mls or less ), draw up 8mls into your syringe, then let sit to cool for a few minutes. Then while it's warm but not hot, suck up 2ml's of karo. Shake really good, and allow to cool a little longer. Inject this forcefully into the LC ( make certain the contents of the syringe isn't too hot (that'll hurt/kill the myc!), then shake the hell out of the LC jar, so as to mix the karo thoroughly.

Now you know there's plenty of oxygen and plenty of nutes. Throw back into incubator and wait another few days... if you don't see anything after 3-4 days, you can assume you just somehow got some bad luck somewhere along the line, perhaps the spore solution that went into that bottle was a pocket of low spore content; did you vigorously shake the syringe everytime you inoculated into an LC?

The final thing to do after both the above, if you're really wanting to experiment and see what may have been the problem, is to go ahead and inject 1/2cc - 1cc spore solution into the jar. Give it another week and see what goes on. Otherwise, chuck it and call it quits. I've had more than one LC jar just inexplicably fail to do anything - frustrating! I've also managed to save a couple doing the above procedure ( I've also killed one w/ too much karo! ).

Anyhow... kinda lenghty and maybe more trouble than you feel like puting into one lousy LC jar! <grin> Just remember to try the aeration first, because it's possible that's the only problem; you don't want to risk puting in too much karo unless the other changes proved unsuccessful.

Best of luck!


--------------------
Nature is the Technology of the Divine.


Edited by VALIS (05/23/05 04:06 PM)


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