bipin dalmia dalmiabk at
Fri Dec 9 17:38:26 EST 1994

In article <stauth-081294142709 at> Darren Stauth,
stauth at writes:
>We recently started making competent cells using DMSO and PEG3350 and we
>were excited at its ease and efficiency.  We are using DH5-alpha cells
>transforming TA ligations.  However, we are now running into the problem
>cell lawns appearing on our transformation plates containing amp.  First
>the transformants appear and then a few hours later the lawn becomes
>visible.  We have tested the amp and the cells both are fine.  Is the
>treating of the cells to make them competent somehow causing them to be
>resistant to amp?  Has anybody experienced this problem?   Any

the 'satellite' colony effect that you are seeing is due to the
ampicillin around the true transformants getting degraded by some
secreted b-lactamase and/or inactivated by the lowering of pH when cells
grow. you may try higher amp. concentration (upto 200 ug/mL) or switch to
the more expensive carbenicillin. carbenicillin is less sensitive to low
pH and in my experience never gives satellite colonies. use carbenicillin
at 100 ug/mL.


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