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OfflineNightPuma1
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Question about inoculating agar..
    #24580037 - 08/26/17 12:35 AM (3 years, 8 months ago)

I had a question about inoculating agar from syringe.

I have read that you are only supposed to put one drop from a flame sterilized syringe into the center of a sterile agar petri dish. I have also read that this droplet is not supposed to roll around as that will spread the contams found in the droplet around on the agar.

How would it ever be possible, then, to wrap a petri dish in parafilm after inoculating with one drop without letting the drop roll around on the agar? Seems like it would be pretty hard to keep completely level while wrapping it up... especially with arms awkwardly shoved into a SAB and in gloves.

Also, what happens if I don't wrap the petri dish in parafilm?


Can I just leave all the petris in my SAB when finished?


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OfflineWeavieWonder
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Re: Question about inoculating agar.. [Re: NightPuma1]
    #24580085 - 08/26/17 01:05 AM (3 years, 8 months ago)

Quote:


How would it ever be possible, then, to wrap a petri dish in parafilm after inoculating with one drop without letting the drop roll around on the agar?




Elevate the dish (a wire rack works great), and then spin it as you wrap it, leaving it flat on the surface.

Quote:

what happens if I don't wrap the petri dish in parafilm?




The culture will be wide open to contaminants.  Wrap em up, always.

Quote:

Can I just leave all the petris in my SAB when finished?




That's what I would do.  Don't move the petris until the spores germinate.


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OfflineNightPuma1
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Re: Question about inoculating agar.. [Re: WeavieWonder]
    #24580132 - 08/26/17 01:41 AM (3 years, 8 months ago)

So if I tilt at all should I trash the petri dish?

Ok another problem:

After I pressure cook my agar in a glass whiskey bottle for 45 minutes, it usually takes around 3 hours to cool to a pouring temperature and typically I can't wait that long and have to come back the next day.

How to best heat it back up to pour it into the petri dishes? I've tried the microwave with marginal success...

Also, what should I cover the mouth of the whiskey bottle with while it is in the PC and microwave? Something that breathes but won't let contams in...


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OfflineWeavieWonder
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Re: Question about inoculating agar.. [Re: NightPuma1]
    #24580209 - 08/26/17 02:49 AM (3 years, 8 months ago)

Just try the best you can to keep them level.  If it runs a little, it runs. Not a huge deal.  No, don't trash it.

Take your agar media bottle(s) out of the PC after the pressure drops to zero. They will be cool enough to pour in about an hour.

If you can't get to it all the same day, then leave your bottles in the PC.  Reheat them in the PC when you're ready to pour.  Cover the tops of your bottles with foil before PCing.  I like to have some sort of filter.  If you want, drill a 1/4" hole in the cap, and cut a piece of SFD to put inside of it.

Check out c10's agar guide in my signature.


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InvisibleAlexSmitt
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Re: Question about inoculating agar.. [Re: WeavieWonder]
    #24580435 - 08/26/17 07:55 AM (3 years, 8 months ago)

I've never used the microwave but I read you have to microwave it a little, shake it, a little more at half power. I wouldn't put anything over it, I'd open the cap a quater turn and see how that works. As for the original question, since it's most likely an ms syringe you're talking about then I'd try to move it around, it's going to get cleaned up anyway so more areas growing gives you more options to choose from rather than all your spors and any contams germinating in the same spot.


--------------------
:tardpig: it's only logical:spock:


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OfflineMirlin
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Re: Question about inoculating agar.. [Re: AlexSmitt]
    #24580450 - 08/26/17 08:08 AM (3 years, 8 months ago)

Quote:

AlexSmitt said:
I've never used the microwave but I read you have to microwave it a little, shake it, a little more at half power. I wouldn't put anything over it, I'd open the cap a quarter turn and see how that works. As for the original question, since it's most likely an ms syringe you're talking about then I'd try to move it around, it's going to get cleaned up anyway so more areas growing gives you more options to choose from rather than all your spores and any contams germinating in the same spot.




The worst advice i had ever given was sterilizing anything with a microwave...just dont.

As far as a dirty syringe, it does not matter if the solution spreads, thats why you re cleaning it up on agar.

Take the wedge from the cleanest portion on the plate and in two to three transfers it will be contam free. If the solution stands in one spot it will take more transfers to isolate out a clean culture due to the proximity of the contaminated spores germinating along side the (assuming) p.cubes.

I should have read alexsmiths post better... hes saying it how it is.


Edited by Mirlin (08/26/17 08:09 AM)


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InvisibleShroomway
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Re: Question about inoculating agar.. [Re: Mirlin]
    #24580551 - 08/26/17 09:49 AM (3 years, 8 months ago)

I made a depression right in the middle of my petris with the flame sterilized back of my scalpel first. Then I put a drop of spore solution there. It doesn't run out. Ideal solution.

From BOD's easy agar TEK I learned that you can use a water bath that is 119F/48C to cool the media bottle down faster. Take water that is 119F/48C and put the bottle in. The water will heat up some. Wait for it to cool down again till 119/48C and poor immediately.

If agar is left in your bottle you can use the microwave to reheat as you said with shaking in between. It needs to reach 180F/82C to become liquid again. Never walk away cause it can cook without warning... I am talking from experience :wink:. I do flame sterilize my glass media bottle very quickly before I pour. But with the cap screwed on so contaminants can be blown into the mouth of the bottle

If you want to avoid condensation you can sterilize your glass petris in the oven so you know for sure they stay dry. You can do it a day before if you like


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