This study aims at development of an approach for selection of strain, which has capability for oxidation of broad-range of chloro-substitute phenols. A multiplex PCR was optimized targeting loci involved in phenol and chlorophenol degradation, which was used to select activated sludge samples and also to assess the degradative genotype of isolates. The isolated strains were screened on the basis of RAPD analysis. In parallel, physiological experiments were carried out with activated sludge samples and isolated bacteria by respirometric analysis. Based on cluster analysis of RAPD pattern and respirometric data, the isolate G20 was selected and identified by using 16S rDNA sequence analysis as Citrobacter freundii strain HPC255. The strain could oxidize different substituted chlorophenol molecules. Such strains could provide the pool of intermediates, which can further be degraded by the associated population, thus helping in maintaining the synergistic association of catabolic activity in activated sludge.