Epitope Tagging Suggestions Wanted

Arle Kruckeberg arle at biovax.leeds.ac.uk
Tue Feb 27 18:41:04 EST 1996


In article <4gpjmb$dia at winx03.informatik.uni-wuerzburg.de>
krasel at wpxx02.toxi.uni-wuerzburg.de (Cornelius Krasel) writes:

> Dr. Arthur L. Kruckeberg (bmbckh at biovax.leeds.ac.uk) wrote:
> [as a reply to a question which epitope tag to use?]
>  
> > Why not try the Strep-tag from Biometra. The tag is small and designed

I didn't write this! It was Chris Hoyle, who hasn't reconfigured the
news-reader to his own name...

I have had good success with the phage T4 gene 10 epitope at the
C-terminus of a membrane protein. The peptide is of neutral charge, so
doesn't break von Heijne's Positive Inside rule (HA, FLAG and c-myc are
negatively charged), it works away from the N-terminus even though it
is from the N-terminus of the gene 10 protein, and for 2 proteins, the
tagged version is 'fully' functional. The monoclonal doesn't light
anything else up in a Saccharomyces cerevisiae crude membrane prep, on
a Western. Haven't tried IP or IF yet, nor compared signal strength of
multiple vs single tags. The monoclonal is available from Novagen, the
epitope codons were introduced by overlap-extension PCR.

Not associated with Novagen, though they kindly gave me a free sample
to try on yeast extracts.

Arle
*************************************************
a.k.a. Dr. Arthur L. Kruckeberg
 Department of Biochemistry and Molecular Biology
University of Leeds        phone  +44 +113 2333172
Leeds LS2 9JT              FAX    +44 +113 2333167
Great Britain              arle at biovax.leeds.ac.uk
*************************************************



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