Team:Virginia United

We used a co-design approach to construct a multiple-compound biosensor that can detect heavy metals (arsenic, mercury, copper) in aquatic environments. The design uses a logic system on three different regulatory levels of the cell. One of the approaches utilizes the operator sites of regulatory promoters, hybridizing two promoters’ operator sites into a single co-sensing promoter. In order for the hybrid promoter to initiate transcription, two target metals that control the operator sites must be present. Since, the hybrid promoters are attached to a single fluorescent protein, the detection of both metals can be measured using fluorescence. The second approach utilizes a fluorescent protein complementation system. When a target metal is detected by a cell, it will transcribe a portion of the non-fluorescent protein. Upon translation, the portions of the fluorescent proteins will bond together and fluoresce, reporting the presence of the two target metals. The third approach allows each of the sensory reporters to express a fluorescent protein in the presence of its target metal. If multiple target metals are detected by a culture, fluorescence spectroscopy is used to separate out the wavelengths of each fluorescent protein, which then determines what compounds are present in the system.

In all three designs we are amplifying the signal that each E. coli cell emits once it is exposed to the target compound with a quorum sensing system. This is because in small doses each metal may not individually be toxic to fish. Each cell releases a signal when exposed to the target compound, which is then recognized by neighboring cells. A fluorescent protein is attached to the promoter that recognizes the signal, establishing a more rapid, binary-like response time in the system. The overall goal of the project is to create a set of interchangeable inputs and outputs serving a wide variety of applications such as in bioremediation machinery.