favorite expression vector

Michael L. Sullivan mlsulliv at facstaff.wisc.edu
Thu Oct 23 09:40:13 EST 2003

<x-flowed>If you're primarily interested in making protein for anybodies, I
think lots of vectors will suit your needs since you don't
necessarily need to worry about whether the protein will be soluble
or not.  [Actually, if your protein is insoluble, that could be a
plus in terms of purifying.]

One I like is pET28a.  This has kan as the selectable marker, so you
don't need to worry so much about losing selection like you do  with
amp.  Depending on what enzyme you choose to do you're cloning, you
can make the expressed protein either with a his-tag or without.

One other consideration is choice of cells for doing the expression.
I have found that for some of the plant proteins I have worked with,
using codon optimized cells [like BL21(DE3) RIL] can make at least a
modest difference in the level of protein expression.  I've seen 2-3
fold higher expression in the RIL cells, but I know some have seen
even bigger increases.

I hope this helps.


PS-  I have no affiliation with Novagen or Stratagene (a source of
the RIL cells)

>I would like to know if anyone can recommend (or
>not) an expression vector. We have a gene encoding
>a membrane protein that we want to make antibodies
>to. Any suggestions? pETs?  Thanks,in advance, Sandy
>- - - - - - - - - - - - - - - - - - - - -
>Sandra Berry-Lowe
>Associate Professor
>Department of Biology
>University of Colorado at Colorado Springs
>tel: 719-339-5276
>fax: 719-262-3047
>slblowe at uccs.edu
>- - - - - - - - - - - - - - - - - - - - -


Michael L. Sullivan, Ph.D
Research Plant Molecular
U.S. Dairy Forage Research Center
1925 Linden Drive West
Madison WI, 53706

(608) 264-5397 Phone
(608) 264-5147 Fax


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