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Agar won't solidify properly: a conundrum


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#1 elabrant

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Posted 09 February 2014 - 04:47 PM

Hello,

I am having some trouble with my MS agar medium. It's 1/2x MS with 6 g/L agar as the gelling agent. I usually pre-melt the agar and mix the pre-melted agar with the nutrient solution before dispensing into 25 mm test tubes. The medium will solidify if left at room temperature BEFORE pressure cooking, but after pressure cooking (30 min @ 15 psi) it remains a viscous liquid that never sets into a solid gel. I've tried a number of ways to make the medium, including tests of pre-melting agar vs. melting in the pressure cooker (both of which solidified). I can't find anything online about agar that won't solidify properly. My only guess is maybe the pH isn't perfect or something in the MS salts/vit mix is keeping it from solidifying, but that shouldn't cause any problems as MS media obviously should solidify. Any ideas?
 
Cheers!
 
I should also add that I had the same issue when trying Gelzan. I mixed 2, 4, and 6 grams into 500 mL batches of ddH2O and pressure cooked for 30 minutes @ 15 psi and none solidified. 


#2 El Crazy Xabi

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Posted 09 February 2014 - 05:04 PM

Check your pH

 

pH starts to affect to agar if it is autoclaved for 15 min pH=3

 

Gellan gum solidification at not-extreme pH is dependant on divalent cations, e.g. Mg, and usually needed to be added as supplement for gelling. At pH 2 or below it doesn't require this and gels without the addition of any extra stuff

 

Wait 6g/L agar? That's 0.6%, that's very low agar concentration. Any agar bottle usually says the recommended minimum concentration for gelling. Likely the same for the gellan gum






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