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Distributed Sensor Network

Distributed Sensor Network. Graham Wiley Leonid Sukharnikov. Levels of Abstraction - System. HSL Amplifier. Arabinose Sensor. Levels of Abstraction - Devices. HSL Sender. Arabinose Promoter. HSL Sender/ Reciever. CFP. YFP. Levels of Abstraction - Parts. Expected Output.

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Distributed Sensor Network

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  1. Distributed Sensor Network Graham Wiley Leonid Sukharnikov

  2. Levels of Abstraction - System HSL Amplifier Arabinose Sensor

  3. Levels of Abstraction - Devices HSL Sender Arabinose Promoter HSL Sender/ Reciever CFP YFP

  4. Levels of Abstraction - Parts

  5. Expected Output Media plate w/ bacterial lawn Expanding YFP ring surrounding CFP circle shows signal propogation

  6. Mechanism Arabinose

  7. Mechanism HSL

  8. Mechanism HSL HSL

  9. Construction 1 2 3 4 parts Final

  10. Construction Results 3 2 1 4 parts Final

  11. FUTURE WORK • Check construct via sequencing • Reconstruct if necessary

  12. Detecting small molecule signaling using phosphorylation dependent mechanism in E.coli Simone Macmil Durga P Sarvepalli 4 November 2006

  13. SMALL MOLECULES • Smallest part of pure chemical substance that retains its structure and properties. • Play an important role in multiple signaling mechanisms

  14. APPLICATIONS OF SMALL MOLECULES IN BIOLOGY • Small molecule drugs • Biopolymers • Synthetic peptides • Primers

  15. NEED FOR SMALL MOLECULE DETECTION • Understand molecule – protein interactions • Effect of molecules on the viability of cells • Genetic changes caused by molecules used in molecular therapy • Drug discovery

  16. CURRENTLY AVAILABLE TECHNIQUES • Analytical: NMR, Western blot, Spectrophotometry, Chromatography, ELISA • Small molecule – protein interactions : Nanowire sensors ( Wang et al , PNAS 2005;102;3208-3212) • Disadvantages: whole cells cannot be used • Using micro channels whole cells can be used and less amount of reagents required • Micro channels can also be customized according to the design of the experiment and allows studies to be conducted under flow

  17. FABRICATION OF MICROFLUIDIC CHANNEL Glass chrome Mask Photoresist Substrate Exposure to UV Light Pattern template PDMS PDMS with channel PDMS - Poly dimethyl siloxane

  18. MICROFLUIDIC CHANNEL DIMENSIONS Channel width : 200µm depth : 50 µm

  19. DEVICE Aspartate detector I6110 J04500 C0082 B0015 OmpR LacI Tar- EnvZ receptor EYFP Induction

  20. METHODS Parts from the registry Restriction enzyme digestion Sequential ligation to create construct Ligate into pUC Transformation Sequence transformants Grow selected transformants in micro channels

  21. RESULTS Aspartate detection Cells grown in Minimal Media lacking aspartate and induced with IPTG Cells fluoresce after 30 min in the presence of LB Microfluidic channel

  22. RESULTS Movement of E.coli in the channel

  23. FUTURE WORK • Improvise microchannels • - Study flow of bacteria under various conditions - chemotaxis, sensitivity of bacteria in the gut to pH, temperature. • Detect low concentration of small molecules formed during bio-industrial production. • -Design appropriate sensors for detection (small peptides and sugars)

  24. ACKNOWLEDGEMENTS Dr. Bruce Roe Dr. Matthias Nollert Dr. David Schmidtke Dr. Fares Nazar Dr. Randall Hewes Doug White Sandra Bryant

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