Plasmid transfer detection in soil using the inducible λPr system fused to eukaryotic luciferase genes

Abstract

We report a model system for plasmid transfer analysis using the regulated lambda phage right promoter, lPR, fused to luc and lucOR as reporter genes. We have demonstrated that the systems cI857-lPR::luc and cI857-lPR::lucOR are temperature-inducible in Escherichia coli but not in other Gram-negative bacteria analyzed, enabling detection of luminescence when plasmids were mobilized from E. coli to those Gram-negative backgrounds. Using light for the detection, we have observed plasmid transfer from E. coli harboring RK2 and R388 derived plasmids to Pseudomonas putida KT2440 (co-introduced with donors) and to indigenous microorganisms, in vitro and in nonsterile soil microcosms. The importance of nutrients for an efficient plasmid transfer in nonsterile soil microcosms has been confirmed. When plasmid transfer experiments were carried out into nonsterile soil microcosms, significant populations of indigenous transconjugants arose. This system provides efficient marker genes and avoids the use of antibiotics for the selection of transconjugants.