Rearranging plasmids, which strain to use

[Clemens Suter-Crazzolara]

Gabor Keresztes wrote:

> Dear netters,
>
>     My aim is to express two similar proteins (wt and mutant) in
> eukaryotic cells. To this aim I chose the pIRES vector which enables the
> expression of two genes from one plasmid.  However, when trying to put
> the second insert into the vector the resulting clones will invariably
> (16 out of 16 prescreened by PCR)  be rearranged by the time I grow up
> the bacteria (strain DH5alpha)  for minipreps.



> Or should I straigh away forget the idea of using a bicistronic
> vector??
>
>                                         Gabor Keresztes, Ph.D

That' s right: forget it.
yu may even get problems if these two inserts are on different plasmids.

clemens suter-crazzolara, PhD