Isolation and characterization of altered plasmids in mutant strains of Pseudomonas putida NCIB 9816

The ability of P. putida NCIB 9816 to grow with naphthalene (Nah+) and salicylate (Sal+) is correlated with the presence of an 83 kilobase (kb) conjugative plasmid, pDTG1. Derivatives of pDTG1 were obtained from cells after exposure to halogenated analogs of naphthalene or salicylate. The selection of mutants having a Nah-Sal- or a Nah-Sal+ phenotype could be enhanced by the addition of triphenyltetrazolium chloride to the indicator medium. Structurally modified plasmids were characterized by restriction endonuclease digestion and Southern hybridization experiments. The region of pDTG1 DNA that encodes the enzymes responsible for the conversion of naphthalene to salicylate was identified. The structural changes in mutant plasmids were correlated with the absence of essential enzymatic activities.