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Offlinejoehinkle
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Shake agar prior to Solidifying
    #10200265 - 04/20/09 08:23 AM (14 years, 11 months ago)

I bought some premixed agar from FP.

When test tubes (agar + wood) for slants comes out of the PC, the mixture is separated with some sediment on the bottom.

The first set I made, I shook prior to letting them solidify but ended with agar bubbles populating the top to the slant.

The second set I made I did not shake and left the sediment at the bottom.

When using agar, should they be shaken just before allowing them to solidify and live with the bubbles?

Edited by joehinkle (04/20/09 08:33 AM)

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Offlinesolumvita
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Re: Shake agar prior to Solidifying [Re: joehinkle]
    #10201025 - 04/20/09 11:04 AM (14 years, 11 months ago)

Are you dissolving the agar in boiling water before you put it into the PC?


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Invisiblelipa


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Re: Shake agar prior to Solidifying [Re: joehinkle]
    #10201102 - 04/20/09 11:20 AM (14 years, 11 months ago)

I just twirl them a little when I set them down to solidify. They are  usually pretty darn hot still when I do that. A little sediment is ok. I cant believe you are shaking culture tubes with agar in them. By bubbles you mean a bunch of agar all over the side of the tube. Like you can't see inside them.

Lipa

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Offlinejoehinkle
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Re: Shake agar prior to Solidifying [Re: lipa]
    #10201383 - 04/20/09 12:34 PM (14 years, 11 months ago)

The directions on FP's agar does not state that the agar needs to be boiled prior to placing into a permanent container - it only states to PC it.

RR's video does not boil prior to pouring into test tubes.

I searched the forums and got a mixture of suggestions to boil and not to boil.

I was concerned about the sediment with the first set of bubes so I shook it after removing it from the PC.  Still hot - but it formed bubbles.

Here is a pic - I have already seeded it with a slice of agar mycelium.



I have 7 more cultures to add to slants so last night I made up 40 new tubes.

I did not shake this time - since RR did not shake them on his video.

The pic below is the worst of the lot as far as sediment.  My only concern is the composition of the agar mix for extending the life of any culture I place it them.



Before I seed these 40 slants with mycelium (5 slants per mushroom culture) I just want to make sure I'm not starting out with garbage.

Thanks in advance for your comments.

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InvisibleJef
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Re: Shake agar prior to Solidifying [Re: joehinkle]
    #10203836 - 04/20/09 08:35 PM (14 years, 11 months ago)

I always boiled. I did MEA.

I always had sediment too, and consider that normal.  Of course the last plates had more sediment than the first.

There was no performance difference that I could discern.

I think you're all okay.


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Offlinejoehinkle
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Re: Shake agar prior to Solidifying [Re: Jef]
    #10208884 - 04/21/09 05:45 PM (14 years, 11 months ago)

Lesson Learned.

The unshaken slants looked OK ... so I setup the flow hood and started to do some transfers from dish to slant.

As I placed a sample in the slant I noticed a small amount of liquid.

I looked at all 40 slants and 80% have liquid which means they did not cure correctly ....

Now I'm disposing of all 40 slants and starting over.

I did not know you could get a partial solidification with agar ...

That sediment on the bottom - (I shook and mixed with the first set of slants and they came out ok) must be part of the solidification reaction.

Hard lesson to learn (time spent wise)!

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OfflineRogerRabbitM
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Re: Shake agar prior to Solidifying [Re: joehinkle]
    #10210253 - 04/21/09 09:02 PM (14 years, 11 months ago)

I always shake the bottle like hell prior to sterilization and before filling the slants.  That tends to reduce the sediment.  I also make up the FP agar about 20% weaker than called for in the instructions.  This takes care of most of the sediment and the slants solidify normally.

When pouring plates, I always give the bottle of agar a good swirl just before pouring.  It also helps to let the agar cool nearly to the point it's starting to solidify before swirling and pouring.
RR


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Offlinesolumvita
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Re: Shake agar prior to Solidifying [Re: joehinkle]
    #10211206 - 04/21/09 11:44 PM (14 years, 11 months ago)

Quote:

joehinkle said:

As I placed a sample in the slant I noticed a small amount of liquid.

I looked at all 40 slants and 80% have liquid which means they did not cure correctly ....





most likely you poured the agar when it was too hot, that causes condensation and accounts for the water in the tubes, only time i get partial solidification is when i make "soft water" agar and that is on purpose.


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InvisibleJef
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Re: Shake agar prior to Solidifying [Re: solumvita]
    #10211241 - 04/21/09 11:51 PM (14 years, 11 months ago)

If solumvita is correct, you can just pour off the excess.


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Tell me and I will forget. Show me and I will remember.  Involve me and I will learn.

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Offlinesolumvita
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Re: Shake agar prior to Solidifying [Re: Jef]
    #10211961 - 04/22/09 03:19 AM (14 years, 11 months ago)

that's exactly what I do


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One of these days all the answers will be revealed until then we learn from each other!

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Offlinejoehinkle
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Re: Shake agar prior to Solidifying [Re: solumvita]
    #10214740 - 04/22/09 05:00 PM (14 years, 11 months ago)

As it turned out - there was NOTHING wrong with the slants.

It was condensation I was seeing.

Live and learn by our mistakes!

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