pET expression help needed

Tony Meyn Tmeyn at ccit.arizona.edu
Fri Jul 22 13:41:14 EST 1994


In article <30of4n$cks at mserv1.dl.ac.uk>, Clemens Suter-Crazzolara
<un691cs at genius.embnet.dkfz-heidelberg.de> wrote:

> > 
> > In your posting you write:
> > >
> > >hello, dear colleagues,
> > >
> > >is it possible to put the t7 tag used in the pET system at THE C-terminus 
> > >of a protein ? is it then still recognized by the corresponding antibody ?
> > >i noticed that novagen offers the HIS TAG both at the N- and C- terminus,
> > >but the T7 tag only at the N-terminus.
> > >
> > >thanks, clemens
> > >    
> > >
> > 
> > T7 isn't a tag. It's a promoter element recognized by T7 RNA polymerase
> > which 
> > is essential for transcription. 
> > 

T7.Tag is a system used by Novagen for the purification of proteins 
expressed in some of the pET vectors.  Genes cloned into the MCS  of
these vectors will contain a 12 amino acid fusion at the N-terminus.  
The fusion is the natural N terminal end of the T7 major capsid
protein.  

This fusion is an epitope for the T7.tag antibody.  Thus,
immunoprecipitation
can be used for purification of the recombinant protein.  As to your
orginal question, I have no idea if the fusion will be equally efective at
the C terminus of your protein.  I would call Novagen and ask them
directly or send them a note.

Novagen, Inc.
597 Science Dr.
Madison, WI 53711 USA
800-526-7319
608-238-6110

Tony Meyn
University of Arizona



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