How to break out bacteria on the plate

Bernard Murray bernard at elsie.nci.nih.gov
Wed Jun 12 03:07:01 EST 1996


In article <31BE2456.2967 at bioneer.kaist.ac.kr>, jhahn at bioneer.kaist.ac.kr 
says...
>
>I'm now finding the method to detect a enzyme activity on agar plate.
>It is intracellular enzyme and its activity can show halo on the plate.
>If we can break out our bacteria on the agar plate as it is,
>we can find colonies that have the enzyme.
>Is there any method or reagent to break out bacteria on the plate?

How about lifting colonies onto a membrane and then breaking the bacteria
there?  You can then treat the bacteria aggressively enough to ensure
that you release the enzyme yet you would also be confident that you
can go back and collect live bugs from the positive colonies.  If
diffusion is a problem then you can make a real replica plate or perhaps
overlay the membrane with agarose/agar etc.  Most molecular biology
methods texts will give recipes for colony lifts and for replica plating.
	If your bacteria are hardy enough you may be able to include
an agent that increases permeability in the agar of your initial plate
- I'm thinking of something along the lines of cholic acid or deoxycholate
as used in selective bacterial media.  You'd have to do some initial
tests just to see what reagents and concentrations you could use.
	I hope that this helps,
		Bernard

Bernard Murray, Ph.D.
bernard at elsie.nci.nih.gov  (National Cancer Institute, NIH, Bethesda MD, USA)
"If I knew what was going on I'd really be indignant." Crow T Robot, MST3k	
				(City Limits)




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