United States of America As Represented by the Secretary of the Air Force

The University of Tennessee Research Foundation

James T Fleming

Gary S Sayler

Michael Leon Simpson

Bruce Michael Applegate

A method for cell-based combinatorial logic includes the steps of providing at least one genetically engineered cell, the genetically engineered cell having at least one transcriptional unit. The transcriptional unit includes a gene and a promoter, wherein application of a stimulus to the promoter results in the expression of a gene product. An energetic or chemical stimulus is applied to activate the promoter, wherein the detection of an output signal corresponds to the presence of a gene product. The cell can include a plurality of transcriptional units configured to form logic gates. The logic gates of a plurality of cells can be operably interconnected by release of output signals, such as chemical stimuli.

Methods for cell-based combinatorial logic

The differential display (DD) technique, widely used previously for eukaryotic gene discovery, is optimized to detect differential mRNA transcription (an expressed gene) from both pure culture and soil derived bacterial RNA. A model system using toluene induction of TodC1 in Pseudomonas putida F1 is used to optimize the procedure. Once optimized, an arbitrary primer for the RT step in conjunction with the same arbitrary primer and a Shine-Dalgarno (SD) primer for the PCR reaction is used to detect the expressed gene. The invention thus provides a method for discovery and acquisition of novel genes from environmental microbial communities that avoids the traditional steps and inherent bias due to the culturing of environmental isolates.

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Knoxville, TN, United States of America

James T Fleming