cotransfect or bicistronic

Rizwan Haq haqr at oci.utoronto.ca
Mon Apr 10 21:21:34 EST 2000


You can get the IRES puro/neo/hygro vectors from Clontech.

Rizwan

bradley turner wrote:

> Hello Alistair,
>
> I hate to ask this, but do you know of a reference or a supplier
> for the 'IRESpuro' vector? It sounds interesting!
> Does this vector allow the expression of two genes
> simultaneously from a single vector?
>
> Thanks,
> Brad Turner
>
> ****************************************************************
>                     Bradley Turner
>                 Beth Israel Deaconess
>                     Medical Center
>
> Harvard Medical School          617-667-1215 phone
> Division of Gastroenterology    617-667-2767 fax
> Room Dana 536                   bsturner at biosun.harvard.edu
> 330 Brookline Avenue            bturner at caregroup.harvard.edu
> Boston, MA 02215                turner at sprcore.caregroup.harvard.edu
> ****************************************************************
>
> On 15 Mar 2000, Alistair Forrest wrote:
>
> > We've played around with cotransfections and the IRESpuro bicistronic
> > vector.
> >
> > With cotransfections you are right in the assumption that a large
> > percentage of your GFP expressing cells will also express your gene of
> > interest, however not all of them will, you can tip things in your favour
> > by using relatively little EGFP plasmid comapred to your GOI vector. Those
> > that fluoresce are more likely to contain the GOI compared to a
> > transfection with equimolar amounts (maybe try 4:1 or 9:1). This will give
> > you fewer green cells but they are more likely to contain your GOI.
> >
> > Alternatively use the IRES vector and avoid all the fuss. The IRES puro
> > vector can allow you to select for your overexpressors within a day or two
> > (much quicker than G418 or hygro).
> >
> > good luck,
> >       Al
> >
>
> ---





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