Curing E.coli plasmids

Tamas Gaal tgaal at macc.wisc.edu
Mon Mar 20 19:51:22 EST 1995


Duncan at genesys.demon.co.uk (Duncan Clark) wrote:
>
> Hi Folks,
> 
> Has anyone got a good high efficiency method for curing both colE1 
> replicon based plasmids (pBR322 deriv.) and p15A replicon based 
> plasmids (pACYC series). EtBr, Acridine Orange, electroporation 
> (if plasmids can go in they can also come out!) etc.

Try growing your culture in sub-lethal concentration of novobiocin for
6-12 hours. Novobiocin selectively inhibits the replycation of the 
plasmids.
I would try something like 50-100-150-200 microgramm/ml in LB (5-10 ml 
culture).
Plate the culture which still grows fairly well, screen single 
colonies for the loss of the resistance marker.
More than 50% of the cells should loose the plasmid.



	Greetings
					tamas

Tamas Gaal
Univ. of Wisconsin
Dept. of Bacteriology

tgaal at macc.wisc.edu





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