Escherichia coli K-12 OST3410 was isolated previously as a stable cyclohexane-tolerant mutant derived from cyclohexane-sensitive strain JA300. A plasmid which provides cyclohexane tolerance to strain JA300 was isolated from the OST3410 genomic library. Subcloning and sequence analysis showed that the plasmid contained the robA gene, whose gene product was reported to bind specifically to the right border of oriC. We observed that the robA gene on the multicopy plasmid generally increased the organic solvent tolerance of several E. coli strains. We also observed an increase in the organic solvent tolerance of JA300 carrying the lac-robA fusion gene on a low-copy plasmid by isopropyl-beta-D-thiogalactopyranoside induction. Strain JA300 carrying the multicopy robA plasmid also showed an increase in resistance to a number of unrelated antibiotics and heavy metal ions, and the spectrum of resistance was significantly similar to that of the soxS-overexpressing strain.