The dszABC genes from newly reported dibenzothiophene biodesulfurizing bacterium, Gordonia alkanivorans RIPI90A were cloned and sequenced. The overall nucleotide sequence similarity between the dszABC genes of G. alkanivorans RIPI90A and those of Rhodococcus erythropolis IGTS8 and Gordonia nitida were 83.1% and 83.2%, respectively. A gene transfer system for G. alkanivorans RIPI90A was established employing the Escherichia coli–Rhodococcus shuttle vector pRSG43 as suitable cloning vector, resulting in transformation efficiencies up to 1.6×10⁵CFUsμg⁻¹ plasmid DNA. This stable vector was applied to cloning and efficient expression of the dsz genes under the control of lac promoter. The recombinant strain was able to desulfurize dibenzothiophene in the presence of inorganic sulfate and sulfur-containing amino acids. The maximum desulfurization activity by recombinant resting cells (131.8μM2-hydroxybiphenylg_{dry cell weight}^-1h^-1) was increased 2.67-fold in comparison to the highest desulfurization activity of native resting cells.