recovery of proteins from phenol

Bruce Roe broe at aardvark.ucs.uoknor.edu
Fri Dec 11 08:13:00 EST 1992


In article <1992Dec11.093922.2189 at gserv1.dl.ac.uk>, vioque at cica.es (Agustin Vioque) writes...
>I have a highly purified protein that I want to silver stain on a gel. The
>protein is contaminated with a large amount of RNA which will also silver
>stain. Toget rid of the RNA I have tried a treatment with NaOH (10', 80C,
>0.1 N) but a get a smear of partially degraded RNA upon staining of the gel.
>I am thinking of doing a phenol extraction, but how can I recover the protein
>from the phenol phase in a way suitable for SDS-PAGE? If anybody has an
>idea I would appreciate it very much. Sorry for my poor english.
>A. Vioque
>vioque at cica.es

Hi,
	Have you though about or tried RNAseA and/or RNAse T1 to degrade
 the RNA?  A trace amount of either or both enzymes often is used to
 remove RNA contamination in plasmid preparations and may be applicable
 to your work.
	As for recovering the protein after phenol extraction, that could
 be extremely difficult if you want any activity, but you can back-extract
 protein from the phenol layer and maybe see enough on a gel.  I have no
 experience with this but check out the literature for RNAse T1 purification.
 As I recall the purification of T1 has a phenol extraction step and a back
 extraction that may give some insite into a phenol-based cleanup of your
 protein.

 Cheers........bruce
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