Single-Cell Fusion PCR for Biocontainment Surveillance

Engineered microorganisms hold great potential for shaping complex industrial and environmental processes. However, introducing genetically modified organisms into an environment always holds the potential for horizonal gene transfer, thus requiring strong biocontainment strategies. While significant energy has been directed towards biocontainment, it is currently difficult to assess the effectiveness of these methods. Common molecular methods such as short- and long-read sequencing of microbiomes cannot always identify the hosts of rare or extrachromosomal genes. One-step Isolation and Lysis PCR (OIL-PCR), which molecularly links any gene of interest with the bacterial 16S rRNA gene via fusion PCR performed within an emulsion, is an ideal method to assay the effectiveness of biocontainment strategies. OIL-PCR is highly sensitive and has been validated to accurately associate plasmid encoded genes with the bacterial hosts in a complex stool community with up to 99% accuracy.