How to show that the proteins are really labeled?

Richard Kerr kerrr at CRYPTIC.RCH.UNIMELB.EDU.AU
Tue Jul 29 23:40:51 EST 1997


At 20:10 28/07/97 -0500, you wrote:
>Please excuse me if these sound like stupid questions; I don't normally do
>this kind of work.
>
>We have fed a radiolabeled polymer to some bacteria and then did an
>extraction/fractionation on the culture.  One of the fractions is a phenol
>solution that supposedly contains proteins.
>
>What we want to know is whether the bacteria incorporated any of the
>radiolabel into their proteins.  The extract contains 5564 dpm/ml of C-14,
>so there is a good chance that there is labeled protein in there.  However,
>how can we be sure that the counts are coming from protein and not from
>something else?  Is there a simple procedure for getting pure protein out
>of the phenol extract? 

3 volumes of cold (-20 Celsius) acetone overnight at 4 degrees celsius.???
I don't know if it will do something crazy to the phenol ...try it on a
small aliquot first.  Precipitate should be a fine flaky/powdery mass that
you can spin down easily in a benchtop centrifuge.  A note of
caution....acetone dissolves some plastics (yes, I know that phenol isn't
nice either) Falcon tubes or equivalent should be okay..

good luck

 We don't care which proteins are labeled so,
>obviously, anything that separates the proteins from each other would be
>overkill.  Any ideas?
>
Richard Kerr.
The Murdoch Institute,
R.C.H. Flemington Rd, Parkville, 3052,
AUSTRALIA.
kerrr at cryptic.rch.unimelb.edu.au
Phone (61) 3 9345 5045.
FAX   (61) 3 9348 1391.
'The most interesting things about vertebrates occur in the neural crest.'
	Peter Thorogood.




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