GeneGuard: a Modular Plasmid System Designed for Biosafety

Synthetic biology applications in biosensing, bioremediation and biomining envision use of engineered microbes beyond a contained laboratory. Deployment of such microbes in the environment raises concerns of unchecked cellular proliferation or unwanted spread of synthetic genes. To introduce security into bacterial synthetic biology, we have taken on the task of completely reformatting plasmids to be dependent on their intended host strain and inherently disadvantageous for others. Using conditional origins of replication, rich-media compatible auxotrophies, and toxin-antitoxin pairs we constructed a mutually-dependent host-plasmid platform, called GeneGuard. These devices, in isolation and in concert, severely reduce unintentional plasmid propagation in E. coli and B. subtilis, and do not disrupt their intended E. coli host’s growth dynamics. Using a fluorescence-based assay we also show that they limit horizontal gene transfer from hosts to other bacteria and they work as efficient vectors for heavy-metal biosensors.