Leonard Lab

Research

Our lab is developing technologies for inducibly and tunably controlling the lateral flow of genetic information between cells within an evolving population. When combined with the integron machinery (see figure), this system will enable the efficient assembly and dynamic diversification of complex multigenic functions (such as exogenous metabolic pathways). For example, the inducible exchange of engineered shuttle plasmids provides a mechanism by which individual genes and regulatory elements, or even arrays of genes, may be exchanged between cells. In this way, an evolving microbial population may sample a greater range of genetic diversity than could possibly be achieved by simply creating massive libraries and then conducting screening.

 

We are engineering two lateral gene transfer systems based upon two distinct mechanisms for natural genetic exchange: (a) conjugation (based upon the F plasmid tra operon) and (b) transduction by non-lysogenic bacteriophage (based upon the M13 bacteriophage). Each mechanism provides a unique modality of genetic exchange; conjugation is a single cell-to-cell transfer event, whereas phage transduction disperses genetic information much more broadly through a population. Although each of the systems on which our technology is based is well-studied, we currently lack tools for inducibly and tunably controlling these lateral transfer events. Therefore, we have developed several novel genetic regulatory schemes for placing the initiation, coordination, and termination of these transfer processes under the control of an orthogonal signal (such as a small molecule or an engineered transcription factor, which may be tied to an upstream engineered synthetic gene circuit). Our initial experiments have demonstrated the shuttle-mediated transfer of genes encoding various fluorescent proteins, which allows us to quantitatively track gene flow through the population. We are working to quantify the performance characteristics and robustness of these technologies, in part using computational approaches developed through collaborations within this project, as well as integrating the lateral transfer technology with components enabling integron integrase-mediated dynamic assembly and modification of multigenic constructs.

 

These robust platforms for tunable lateral gene transfer will also provide a suite of synthetic biology technologies that should prove useful for both (a) biotechnology applications, including directed evolution or dynamic performance optimization within microbial communities and consortia, and for (b) pursuing fundamental scientific questions related to the structure and dynamics of multicellular microbial networks, such as environmental microbial populations and microfloral communities within the gastrointestinal tracts of multicellular organisms.

 

People

  • Dr. Josh Leonard
  • Mr. Andrew Scarpelli

Related publications

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