RP4 /MU3A-MEDIATED IN-VIVO CLONING AND TRANSFER OF A CHLOROBIPHENYL CATABOLIC PATHWAY/

Chromosomal DNA fragments encoding the ability to utilize biphenyl as sole carbon source (Bph(+)) were mobilized by means of plasmid RP4::Mu 3A from strain JB1 (tentatively identified as Burkhoideria sp.) to Alc aligenes eutrophus CH34 at a frequency of 10(-8) per transferred plasm id. The mobilized DNA integrated into the recipient chromosome or was recovered as catabolic prime plasmids. Three Bph(+) prime plasmids wer e transferred from A. eutrophus to Escherichia coil and back to A. eut rophus without modification of the phenotype. The transferred Bph(+) D NA segments allowed metabolism of biphenyl, 2-, 3- and 4-chlorobipheny l, and diphenylmethane. Genes involved in biphenyl degradation were id entified on the prime plasmids by DNA-DNA hybridization and by gene cl oning. Bph(+) prime plasmids were transferred to Burkholderia cepacia, Pseudomonas aeruginosa, Comamonas testosteroni and A. eutrophus and t he catabolic genes were expressed in those hosts. Transfer of the plas mid to the 3-chlorobenzoate-degrading bacterium Pseudomonas sp. B13 al lowed the recipient to mineralize 3-chlorobiphenyl. Other catabolic pr ime plasmids were obtained from JB1 by selection on m-hydroxybenzoate and tyrosine as carbon sources. 16S rRNA sequence data demonstrated th at the in vivo transfer of bph was achieved between bacteria belonging to two different branches of the beta-Proteobacteria.