Researcher(s)
- Samuel MacDonald, Chemical Engineering, University of Delaware
Faculty Mentor(s)
- Mark Blenner, Department of Chemical Engineering, University of Delaware
Abstract
Engineered microbes are powerful tools with the potential to help solve many modern environmental challenges. However, their uncontrolled release into the environment could disrupt ecosystems. To mitigate this, researchers are focused on developing bio-containment methods that spatially and temporally limit the proliferation of engineered microbes. Leading bio-containment methods include inducible kill switch systems and auxotrophic approaches. For inducible systems, researchers supply an inducer to activate death of an engineered organism, while in auxotrophic systems researchers supply a survival factor that the organism needs to proliferate. These methods are ideal for applications that are spatially limited, such as engineered probiotics, but are impractical for applications that occur over large geographic areas, such as oil spill remediation. The creation of a long-term biological timer addresses this problem by setting an “expiration date” on released microbes, which would allow them to serve their purpose before engineered death occurs. This allows microbes to spread without risk of long-term survival and proliferation into the ecosystem, and can be adjusted to match the necessary duration of their tasks. The timer uses a T7 polymerase fused with a DNA base editor (eMutaT7) to create mutations on a defined DNA segment, which we refer to as a “fuse”. “Roadblocks” in the form of DNA binding proteins or T7 terminator sequences prevent eMutaT7 from progressing downstream on the fuse. However, these roadblocks can gradually be removed through the mutagenizing activity of eMutaT7, which eventually allows the polymerase to reach the downstream portion of the fuse where a killing payload will be stored. The transcription of this payload would result in the death of the cell. The “fuse” has multiple options for tun-ability, such as number of roadblocks and roadblock strength. This timer could be inserted into engineered cells to help protect ecosystems from potential unintended consequences of environmental release.