HA tagged vectors for yeast expression

Ben Lieberman liebe003 at mc.duke.edu
Fri Sep 29 08:16:02 EST 1995


Well, it seems that people are interested in my HA (hemagglutinin) fusion
vectors, since I have been receiving numerous requests to send these
vectors out.  I have been happy to do so and will continue to do so
because I would like to think that science is a cooperative endeavor.  So
if you are interested here is the skinny:

I have built a series of vectors, based on the pRS400 series of yeast
shuttle vectors, with the CUP1 copper inducable promoter and the CYC1
terminator.  Using modular construction (so that you can swap promoters or
Tags if you so desire), I added the HA tag with a S-D sequence and the
start ATG followed by in-frame SSTI and XHOI cloning sites.  This vector
is illustrated:

-BAMHI---CUP1 Promoter----ECORI--SD--MET-HA TAG---SSTI---XHOI-------CYC
terminator---CLAI--

The HA tag is 9 A.A. long and is recognized specificaly by the 12CA5 mAB
(available from Boehinger-Mannheim).  This vector currently comes in three
flavors: integrating, CEN/ARS, and 2 micron, all of which have the TRP
marker.  Since this plasmid is based on the pRS vector series you can swap
in the LEU2 or the URA3 gene using the BSSHII sites in the plasmid.  I
have also constructed a vector with the HA tag fused to the C-terminal,
but this is only in the high copy number flavor.  Again this uses the same
sites and can be swapped with the other markers and replication systems of
the pRS series (isn't modular construction nice?).

To avoid the rush of responses, please be sure that you will really use
this vector before you request it.  I am only one lowly post-doc, and all
responses will be handeled by me alone.  If you wish to share these
plasmids, feel free, but please acknowledge me as the originator.

p.s. Any companies interested in marketing this plasmid series are
encouraged to contact me directly by e-mail

Thanks, hope this helps.

Ben Lieberman, Ph.D.
Duke University

liebe003 at mc.duke.edu



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