Plasmid pSI30 was constructed to increase the sensitivity of detection of a genetically engineered microorganism (GEM) and its recombinant DNA in environmental samples. The broad host-range, mobilizable plasmid contained chlorocatechol (clc) degradative genes, antibiotic resistance genes (ampicillin and kanamycin), and a fragment of eukaryotic DNA. The clc genes encode enzymes that convert 3-chlorocatechol to maleylacetic acid permitting the host, Pseudomonas putida RC-4, to grow on 3-chlorobenzoate. The catabolic phenotype was exploited using enrichment procedures to detect RC-4(pSI30) cells, freeliving in the water column or when irreversibly bound to surfaces. The eukaryotic DNA sequence provided a unique target allowing positive identification by DNA:DNA hybridization. In flow-through microcosms, RC-4(pSI30), undetectable as freeliving cells, was found by enrichment as irreversibly bound sessile forms. These experiments revealed the stability of pSI30 and its utility in a combination detection system for tracking the survival of a GEM and its DNA in environmental samples.