Use of phenotypic suppression for direct selection of loss of aminoglycoside antibiotic resistance determinants

A strain of Escherichia coli containing a conditional drug dependent arginine auxotrophy was used to select for the loss of plasmid and/or transposon encoded kanamycin (Km) or streptomycin (Sm) resistance determinants. Because these determinants inactivate the corresponding drug thus eliminating drug suppression, loss of the drug-resistance determinants was selected directly by growth on minimal media plates containing sub-lethal dosages of the drug. This method was used to select loss of Km or Sm resistance determinants due to loss of plasmids, transposition from plasmid to chromosome, and education of transposons from the chromosome. Drug suppression was compared to phage PRD1 resistance in selecting for loss of plasmid vehicles during transposition and was found to be 10-1,000 times more efficient. Eighty percent of the eductant clones had undergone imprecise eductions suggesting that this method may be useful in selecting stable deletion mutants. An antibiotic suppressible strain of Pseudomonas stutzeri was obtained implying a broad utility of this selection procedure.