This is the first report on alkane degradation in Pseudomonas frederiksbergensis by certain genes cluster of hydroxylase system. The gene encoding alkane hydrolyse of Pseudomonas frederiksbergensis was cloned and characterized. The gene organization in the gene cluster of alkane hydroxylase gene in Pseudomonas frederiksberghensis was a unique one that was different from all known alkane hydroxylases gene families that were found in other alkane degrading bacteria. The alk gene cluster containing homologues to alkane monooxygenase (alkB) was oriented in the same direction, whereas alcohol dehydrogenase (alcDH) was oriented in the opposite direction. The gene encoding alcohol dehydrogenase (alcDH) in Pseudomonas frederiksbergensis was cloned in E. coli and biochemically characterized. It was located in one gene cluster together with alkB on the chromosome of Pseudomonas frderikbergensis. The other genes scattered on the chromosome. Therefore, the study of this gene has been interesting since it has represented a unique phenomenon in genetic engineering and molecular biotechnology to this bacteria.