Inducible eukaryotic expression vector ?
bernard at elsie.nci.nih.gov
Tue Jun 7 18:41:47 EST 1994
In article <2sv5p6$7dp at pobox.csc.fi>, airenne at convex.csc.FI (Kari Airenne) writes:
> Dear colleagues/netters
> I am wondering if any of you have used LacSwitch Inducible Mammalian expressionsystem sold by Stratagene for protein production in eukaryotic cells. As far
> as I have understood right there are only limited number of possibilities for
> inducible protein production in eukaryotic cells and, at least for me, this
> seems to be a good choice for that. So if you have used this system or if you
> know any other good tightly regulated eukaryotic vectors, please let me know.
> Kari Airenne
Ah, a question close to my heart at the moment! I am trying out the LacSwitch
myself. I have transfected CV-1 cells with the lacI expression vector, selected
in hygromycin and am now screening them. Meanwhile I am constructing the
lacO/RSV promoter plasmid. As soon as I have characterised the cells I'll
probably test their switchability with the positive control (CAT) plasmid.
I have yet to hear of anyone actually using this system in anger and the only
criticism I have heard is that it is biased to having the switch OFF so that
good induction can be difficult to obtain. Although you *can* use IPTG in
vivo the half-life is very short (? 30 minutes in mice) so I don't know how
useful this will be for transgenics. I shall keep you posted.
As far as other inducible expression systems are concerned, the other
commercial offerings are based on glucocorticoid induction;
Pharmacia and Clontech offer (?identically) the Lee et. al. (Nature 1981)
version - Pharmacia's is *much* cheaper than Clontech.
USB recently offers the Mader and White (PNAS 1993) version and they also
have it in an EBV form for episomal expression.
There are many useful non-commercial offerings - Wurm et al's heat
shock promoter (PNAS, 1986), McCormick et al's interferon promoter (MCB 1984),
Gossen and co's tetracycline response (PNAS 1992), Richard Hanson's PEPCK
(JBC 1994). Basically, any existing inducible promoter could be a candidate
[ANYONE WANT TO OFFER ANY!]. The "classic", metallothionein, has the major
problem that it is too leaky in the "uninduced" state.
Any other information gratefully received,
ObDisclaimer: No commercial attachments, only buy their products etc. etc.
Bernard Murray, Ph.D.
bernard at elsie.nci.nih.gov (National Cancer Institute, NIH, Bethesda MD, USA)
More information about the Methods