Two previously described methods for gene targeting (replacement) in Escherichia coil were applied to the disruption of the maoCA operon in Klebsiella aerogenes. These techniques involve plasmid-chromosomal integration, resolution of the integrated intermediate and segregation (i) screened using fusaric acid for the counter selection of plasmid replicons carrying the gene for tetracycline resistance, or (ii) by using a temperature sensitive replicon. Both methods were found to be effective to create the desired chromosomal mutants in a Klebsiella strain after some modifications of the original experimental protocols, and may serve as tools for gene targeting studies in Klebsiella and related species.