Here is a puzzler. We have recently had a problem with
satellite colonies on O/N growth of our Amp plates. This
usually happens once a year or so. We have performed all
manner of controls, such as increasing Amp concentration
to 350ug/ml, making new Amp stocks, buying new Amp from
other vendors. We have spread Amp on top of fresh LB
plates (15cm) at varying concentrations (up to the
equivalent of 1 mg/ml!) We still get some satellites,
particularly with high copy number plasmids. We see
satellites whether there are 200 colonies or if there are
10.The number of satellites decreases with increasing Amp
but with certain plasmids, such a pGEX, they still remain.
We don't believe the Amp is being heat-inactivated since
we have taken special care, and the spreading experiments
used fresh Amp. Different people have also been involved
in making plates.We are using concentrations of Amp that
It appears to me that something is inactivating the Amp but
I don't know what.
We are getting somewhat frustrated. The problem will
probably go away soon, but I'm sure it will return again.
Is there something that could be sequestering the Amp? Would
carbenicillin be more effective? Any help would be appreciated.
gayler at immunex.com
The opinions expressed are solely my own.