In article <55t3ph$o6j$1 at montespan.pasteur.fr>
Andre Habel <ahabel at pasteur.fr> writes:
> Hy,
>> I would like to construct a plasmid,which has no antibiotic resistance
> gen. Instead I would like to integrate something else (DNA-sequence,
> small gene, etc.) for convinient selection of positive clones. Due to the
> fact that I want to keep my plasmid as small as possible, the
> selection-marker / sequence should also be as small as possible.
> I would appreciate any ideas/suggestions on that particiular case.
> Plaese e-mail your ideas to ahabel at pasteur.fr>> Thanks in advance :-)
>> Andre
There are a variety of options. Brian Seed developed a plasmid system
some time back called PiVX (no greek letters in email) which uses a
suppressor tRNA gene (supF) as a selectable marker. You transform into
a strain carrying an amber mutation in a seletable gene (like a trpEam)
or you have a strain carrying the plasmid P3 which carries amber
mutants in Amp and Tet. So you can select for those to get
transformants. I don't remember the exact size, but something like
100-200 nt for the gene. The entire plasmid was about 900nt. There are
certainly other tricks you can play with sites etc, like an operator
site for a repressor to cause derepression of an operon. But the PiVX
system is known to work.
Michael Benedik
Department of Biochemical Sciences
University of Houston
benedik at uh.edu
