extremely low copy number plasmid

Alison Leakey leakey at cber.fda.gov
Wed Jan 30 16:43:03 EST 2002


Hello everyone!

I would appreciate any suggestions. I am working with a plasmid that is
in extremely low copy number. Lately we have been trying to increase the
yeild of our preps. Normal mini-preps give us nothing - no visible
plasmd. The QIAGEN maxi prep kit with cultures of up to 2L gives us very
faint bands that are relatively resistant to digestion.  Alkaline lysis
yeilds similar faint bands but also poor quality DNA. We have also
increased the concentration of selective antibiotics in cultures with
only marginal success. We are just about to give up.

Any thoughts??

Alison Leakey

PS. We have used these methods with other low copy plasmids also with
good results so we are not making any obvious mistakes.





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