Hello netters:
I am working in a lab which performs an Agrobacterium tumafaciens
mediated transfection of Arabidopsis plants. In the transformation of
The Agro, an E. coli strain (with the plasmid of choice), as well as a
helper E coli strain (which transfers the DNA from the first E coli
strain to the Agro) are mixed with the Agro (Triparental Mating). Our
problem is this: a successful transformation of the Agro is selected for
by antibiotic resistance, and the Agro is also ketolactose tested; the
strain is then frozen down. When it is used a month or so later, the
strain no longer tests positive (by ketolactose). How can an Agro
strain appear to be switching to E. coli?
We perform multiple selective media streaks, pick single colonies, but
still it persists.
Does anyone have any ideas about what's going on?
Posting to Here or e-mailing me is fine
Thanks in advance
Michael Allen
m3allen at sciborg.uwaterloo.ca
