The bacterial strain Delftia acidovorans P4a, isolated from an extreme environment (heavily contaminated with organochlorines, highly alkaline conditions in an aqueous environment), was found to mineralize 2,4-dichlorophenoxyacetic acid (2,4-D) and 2-methyl-4-chlorophenoxyacetic acid under alkaline conditions. Screening a genomic DNA library of the alkalitolerant strain for 2,4-D genes revealed the presence of the two 2,4-D gene clusters tfdCDEF and tfdC(II)E(II)BKA, tfdR genes being located in the vicinity of each tfd gene cluster. The results showed that the putative genes of the complete 2,4-D degradation pathway are organized in a single genomic unit. Sequence similarities to homologous gene clusters indicate that the individual tfd elements of strain P4a do not share a common origin, but were brought together by recombination events. The entire region is flanked by insertion elements of the IS 1071 and IS 1380 families, forming a transposon-like structure of about 30 kb, of which 28.4 kb were analysed. This element was shown to be located on the bacterial chromosome. The present study provides the first reported case of a chromosomally located catabolic transposon which carries the genes for the complete 2,4-D degradation pathway.