Applied Microbiology and Biotechnology, Vol.82, No.4, 749-756, 2009
A fluorescent, genetically engineered microorganism that degrades organophosphates and commits suicide when required
One way to reduce the potential risk of genetically engineered microorganisms (GEMs) to the environment is to use a containment system that does not interfere with the performance of the GEM until activated. Such a system can be created by inserting a suicide cassette consisting of a toxin-encoding gene controlled by an inducible promoter. We constructed a GEM that can degrade organophosphorus compounds, emit green fluorescence, and commit suicide when required by putting the genes that control these different functions under different promoters. The genes for enhanced green fluorescent protein (EGFP) and organophosphorus hydrolase (OPH) were cloned downstream of the lambda P-L promoter in the plasmid pBV220. These genes could be expressed freely as long as the GEM was metabolizing because the repressor sequence cIts857 had been deleted. The extracellular nuclease gene of Serratia marcescens, without its leader-coding sequence, provided the suicide mechanism. This was put under the control of the T7 promoter to form a suicide cassette activated by the presence of an environmental signal, in this case, arabinose. To improve the reliability of this containment system, the suicide cassette was duplicated within the conditional suicide plasmid. The plasmid carrying the EGFP and OPH fusion genes and that containing the suicide cassette were compatible and coexisted in the same host.