white colonies have no inserts, why?
Dr. Duncan Clark
duncan at nospam.demon.co.uk
Thu Jul 16 11:33:23 EST 1998
In article <9807161054.AA07211 at sun5>, beisz at SUN5.IBP.AC.CN writes
>wo thirds of the transformants are
>white, but only
>5% of these white colonies contain inserts. Who can tell me why?
Trace nuclease in the plasmid prep, RE, RE buffer or any subsequent step
could be blunting the overhangs. Ligation gives an out of reading frame
lacZ hence white colonies. Very very doubtful that intact Pst I and EcoR
I overhangs ligate themselves with T4 DNA ligase.
The problem with being on the cutting edge is that you occasionally get
sliced from time to time....
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