SEGREGATIONAL AND STRUCTURAL INSTABILITY OF RECOMBINANT PLASMID CARRYING GENES FOR NAPHTHALENE DEGRADING PATHWAY

The stability of recombinant plasmid carrying genes for naphthalene mi neralization was determined. A strain of Pseudomonas putida capable of mineralizing naphthalene (Nap(+)) via salicylate (Sal(+)) was isolate d, and all regulatory and structural genes for the whole pathway were found to be encoded on a 25 kb EcoRI fragment of an approximately 83 k b plasmid present in this strain. The 25 kb EroRI fragment was cloned into a tetracycline-resistant (TCR) cloning vector pLAFR3 and the reco mbinant plasmid, pRKJ3 (Nap(+), Sal(+), Tc-R), thus Obtained was trans ferred into the plasmid-free strain Pseudomonas putida KT2442 in order to test the stability of the plasmid. Plasmid pRKJ3 was found to be s egregationally and/or structurally unstable, depending on the growth c onditions. Two types of novel derivative strains having the phenotypes Nap(-), Sal(+), Tc-R and Nap(-), Sal(-), Tc-R with specific deletions of approximately 2 kb and 18 kb, respectively, were obtained.