curing bacteria of plasmids

Pete Muriana MURIANAP at FOODSCI.PURDUE.EDU
Tue Oct 25 10:02:25 EST 1994


In article <38j1b8$me4 at post.its.mcw.edu> ckrepel at post.its.mcw.edu (Candace Krepel) writes:

>I am looking for a reference that would provide a method for curing bacteria
>of their plasmids.  Any help would be appreciated.

>Candy Krepel, Surgical Microbiology Research Lab, Medical College of WI
>ckrepel at post.its.mcw.edu

People have used :
      1. High temperature growth,
      2. Nutrient starvation,
      3. DNA-intercalating dyes,
      4. Novobiocin, naladixic acid, other gyrase inhibitors.

It's best if you have a plasmid-expressed phenotype to follow.  I, and several 
colleagues, have used the Novobiocin method which either works well or not at 
all.  When it worked for 3 people that i know of, we were all using it on 
cryptic plasmids - seemed that it worked so well, that screening of 10-20 
streaked colonies demonstrated that about 30-40% of them lost their plasmid; 
I've tried it to no avail with other plasmids.  We performed growth curves vs. 
control culture at ug/ml levels and used the level that was slightly 
inhibitory to growth compared to the control; perform several consecutive 
transfers with 0.1% inoculum, streak culture, pick 10 or more colonies for 
plasmid screening (or look for loss of phenotype=best if available).  
Novobiocin affects gyrase enzyme which is required for plasmid replication.

Plasmid curing in bacteria.  J.T. Trevors, FEMS Microb. Rev. 32:149, 1986.

Elimination of plasmid pMG110 from E. coli by novobiocin and other inhibitors 
of DNA gyrase.  Hooper, et al., Antimicrob. Agents chemother. 25:586, 1984.

Antagonism of the B subunit of DNA gyrase eliminates plasmid pBR322 and pMG110 
from E. coli.  Wolfson et al., J. Bact. 152:338, 1982.

Two modes of curing transmissible bacterial plasmids.  Salisbury et al., J. 
Gen. Microbiol. 70:443, 1972.

Curing effect of Clorobiocin on E.coli plasmids.  Cejka et al., Mol. Gen. 
Genet. 186:153, 1982.

Curing of E.coli K12 plasmids by coumermycin.  Danilevskaya and Gragerov, Mol. 
Gen. Genet. 178:233, 1980.

Hope this helps. regards, Peter
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*  Peter M. Muriana, Ph.D.             317-494-8284   TEL            *
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