2 genes on one plasmid

Bart Janssen bjanssen at bio.tamu.edu
Thu Nov 16 11:07:28 EST 1995


In article <489j1s$grj at saba.info.ucla.edu>, Sami Kohan <skohan at ucla.edu> wrote:

> I'm trying to build a vector with two genes and was going to use the same
> promoter twice. However I was told that it would cause recomibinations
> and otherwise mess up my experiment, however if the promoters are only
> 50bp it seems to my unexpert opinion to be too small of a homology to
> cause recombination. Any one have any experience with this?

Hi Sami
I've been making binary vectors for plant transformation and several of
these have "double promoters" ranging in size from 300 bp to 2 kbp in both
inverted and direct repeats.  I have never seen any recombination in E.
coli (strain DH5alpha) and have only seen one instance of what could have
been recombination in the Agrobacterium (that was with two 2 kbp promoters
arranged in an inverted repeat).  I'm not even convinced that that
instance was recombination between the promoters.  So I've never had
serious problems with bacterial recombination.

However, I have heard reports from others working with binary vectors who
have made some very unstable "double promoter" constructs.  I suspect that
it very much depends on the individual sequence you replicate.  Obviously
doing your cloning in a rec deficient bacteria doesn't hurt :).

cheers
Bart



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