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From 2012.igem.org

Project Overview

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Arizona State's 2012 iGEM project aims to develop a portable water-borne pathogen biosensor feasible for real-time field application. To achieve both specificity and portability, the team is constructing two biosensors, each capable of detecting a specific pathogenic marker in water-borne bacteria. The first system, a split-enzyme engineered fusion protein, selectively binds to pathogen membranes in water samples and induces a colorimetric response. The second system will detect specific DNA sequences in pathogenic bacteria and activate a similar colorimetric change. The advantage of this design over previous designs in the field lies in the cheap production of probes and the enzymatic chain reaction. In this way, samples can be tested in the field with minimal cost and high sensitivity.

Abstract

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Diarrheic pathogens including E.coli O157:H7 serotype, Campylobacter, Shigella, and Salmonella often contaminate drinking water supplies in developing nations and are responsible for approximately 1.5 million worldwide annual deaths. Current technologies for detection of bacteria include DNA hybridization FRET signaling, electrical detection via immobilized antimicrobial peptides, and PCR amplification followed by gel visualization. Our method of bacterial detection fills a niche in biosensor technology. Our design implies lower costs, higher portability, and a more rapid signal output than most bacterial biosensors. Additionally, our interchangeable DNA probe confers modularity, allowing for a range of bacterial detection. Using a novel split beta-galactosidase complementation assay, we have designed three unique chimeric proteins that recognize and bind to specific pathogenic markers and create a functioning beta-galactosidase enzyme. This functioning enzyme unit then cleaves x-gal and produces a colorimetric output signal. Our research demonstrates success in initial stages of chimeric protein assembly.

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