Mode of access: World Wide Web.

System requirements: Adobe Acrobat Reader.

Part of: Synthesis digital library of engineering and computer science.

Series from website.

Includes bibliographical references.

1. Short history of the use of bacteria for biosensing and bioreporting -- Early warning systems -- Early use of bacterial 'bioreporters' -- References --

2. Genetic engineering concepts -- Introduction to genetic sensing/-reporting circuits -- Central idea -- Intercept design -- Orthogonal design -- Design parts -- Use of transcriptional activators -- Choice of regulatory proteins -- The XylR/DmpR family of transcription activators -- The HbpR system -- Use of transcriptional repressors -- Regulators from heavy metal resistance -- The MerR system -- ArsR-based designs for arsenic detection -- Network interception designs -- General motivation -- SOS response network intercept design -- Promoter engineering -- General notions -- Promoter engineering in the Ars system -- Physiological control of the XylR-regulated Pu promoter -- Dual responsive control switches -- Directed evolution of promoters -- Response heterogeneity in populations -- Engineering new effector specificities -- General concept -- Effector domain mutagenesis in XylR/DmpR-type proteins -- Effector binding pocket modeling -- Aptamers -- Complex signal-transduction chains -- General concept -- Periplasmic binding proteins and phosphotransfer relay -- Multinode networks -- Logic gates, transcriptional noise, amplification -- Reporter proteins -- Choice and specificities of common reporter proteins -- Reporter gene vectors -- Embedding of sensing/-reporting circuits in a cellular chassis -- References --

3. Measuring with bioreporters -- Assay principles -- Relative and absolute measurements -- End-point and kinetic measurements, measurement transformations -- Population measurements -- Spiking -- Theory of analyte provision and transport -- Calculation of compound concentrations in reporter cells -- Non-diffusive and non-conservative bioreporters -- Concept of bioavailability and bioaccessibility -- Bioavailable fractions -- Bioavailability and bioaccessibility reporter measurements -- Bioreporter assay types -- Aqueous assays -- Gas phase measurements -- Solid phase assays -- Method detection limits, accuracy and precision -- References --

4. Epilogue -- Summary -- Future directions -- References --

A. Bacterial bioreporter designs targeting organic compounds -- References -- B. Bacterial bioreporter designs targeting (heavy) metals and metalloids -- References -- C. Bacterial bioreporter designs responsive to toxicity or stress conditions -- References -- D. Example bioreporter protocols -- Quantitative arsenite measurements with an E. coli LuxAB Luciferase bioreporter -- References -- Arsenic measurement using an E. coli GFP biosensor by epifluorescence microscopy -- References -- Arsenic measurements with an E. coli beta-galactoside bioreporter -- References -- Sample pretreatment -- Water -- Rice -- References -- Author's biography.

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Bacterial reporters are live, genetically engineered cells with promising application in bioanalytics. They contain genetic circuitry to produce a cellular sensing element, which detects the target compound and relays the detection to specific synthesis of so-called reporter proteins (the presence or activity of which is easy to quantify). Bioassays with bacterial reporters are a useful complement to chemical analytics because they measure biological responses rather than total chemical concentrations. Simple bacterial reporter assays may also replace more costly chemical methods as a first line sample analysis technique. Recent promising developments integrate bacterial reporter cells with microsystems to produce bacterial biosensors. This lecture presents an in-depth treatment of the synthetic biological design principles of bacterial reporters, the engineering of which started as simple recombinant DNA puzzles, but has now become a more rational approach of choosing and combining sensing, controlling and reporting DNA 'parts'. Several examples of existing bacterial reporter designs and their genetic circuitry will be illustrated. Besides the design principles, the lecture also focuses on the application principles of bacterial reporter assays.A variety of assay formats will be illustrated, and principles of quantification will be dealt with. In addition to this discussion, substantial reference material is supplied in various Annexes.

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