Countries Afgected by Landmines Countries afgected by Landmines - PowerPoint PPT Presentation

e ectrace

Countries Afgected by Landmines Countries afgected by Landmines Image adapted from Handicap International Afgected Countries Non Afgected Countries 2

Conceptual Design Wifj enabled device reads Electrace current and sends data to web app. Read-out from e Electrace μtrace contains E. coli (Landmine) biosensor 3

OUTPUT μTRACE INPUT Electrical Signal Outside the Lab Biosensor - Conductive Curli - Microfmuidic Platform - Landmines - Electron Transport 4

OUTPUT μTRACE INPUT Electrical Signal Outside the Lab Biosensor - Conductive Curli - Microfmuidic Platform - Landmines - Electron Transport 5

Improving the Response BioBrick (BBa_K1316008) ybiJ Promoter 15 100mg/L DNT 50mg/L DNT 0mg/L DNT 10 100mg/L DNT +N 50mg/L DNT +N 0mg/L DNT +N 5 5 5 10 15 20 25 0 Time (hr) Literature: Yagur-Kroll et al ., 2014 6

OUTPUT μTRACE INPUT Electrical Signal Outside the Lab Biosensor - Conductive Curli - Microfmuidic Platform - Landmines - Electron Transport 7

OUTPUT μTRACE INPUT Electrical Signal Outside the Lab Biosensor - Conductive Curli - Microfmuidic Platform - Landmines - Electron Transport 8

Curli in Biofjlm BioBrick (BBa_K1316014) A A A A A A Induction A A A A A A A A B B B Cell Membrane Inducible Constitutive csgB csgA Barnhart and Chapman, 2006 9

Conductive Curli Medium Gold nanoparticle Gold Electrode Substrate Induction Cell without Curli Cell with Curli Chen et al. , 2014 10

Stochastic Curli Modeling Gene Level nm scale Cell Level μm scale Colony Level mm scale 11

Stochastic Curli Modeling Deterministic ODEs Stochastic Worm Like Chains Stochastic Percolation Theory & Graph Theory 12

Percolation 13

Percolation 14

Conclusions from Modeling Induction of conductive Curli results in an electrical signal. This is a qualitative response. 15

OUTPUT μTRACE INPUT e - Electrical Signal Outside the Lab Biosensor - Conductive Curli - Microfmuidic Platform - Landmines - Electron Transport 16

Shewanella oneidensis MR-1 Metal Oxides Extracellular MtrC Space MtrB MtrA Lactate Dehydrogenase CymA Q QH 2 Cytosol Electron Electron Donor Donor Oxidized Adapted From Goldbeck et al., 2013 17

Escherichia coli C43 The fjrst functioning MtrCAB Cluster (BBa_K1316012) Cytochrome C Maturation System (BBa_K1316011) Electrode Extracellular MtrC Space MtrB MtrA Lactate Dehydrogenase NapC Q QH 2 Cytosol Electron Electron Donor Donor Oxidized Adapted From Goldbeck et al., 2013 18

Flux Balance Analysis Determination of Optimal Electron Donor Using COBRA Toolbox Anoxic Electrode Extracellular MtrC Space MtrB MtrA Lactate Dehydrogenase NapC Q QH 2 Cytosol Lactate Pyruvate Adapted From Goldbeck et al., 2013 19

Electron Flow Potentiostat A e - e - Counter Electrode Reference Working Electrode Electrode - MtrCAB + e - Donor Oxidized e - Donor e - e - 20

Custom-designed Bioreactor Nitrogen Gas outlet inlet Sampling tube Silver/Silver Chloride reference electrode Counter Electrode Carbon cloth working electrode Heating mantle Magnetic stirrer-bar 21

Custom-designed Bioreactor Software Development in LabVIEW Experimental Setup Potentiostat Circuit and Software 22

Measurement of Current MtrCAB & Ccm BioBrick (BBa_K1316012 & BBa_K1316011) 10 E. coli C43 + MtrCAB & Ccm 9 E. coli C43 8 7 Normalized Currnet [mA] 6 5 4 3 A 2 e - e - 1 - + 0 1 2 3 4 e - Time [hr] e - e - 23

OUTPUT μTRACE INPUT Electrical Signal Outside the Lab Biosensor - Conductive Curli - Microfmuidic Platform - Landmines - Electron Transport 24

Microbial Biosensors Now: Numerous but Impractical 25

Microbial Biosensors Now: Numerous but Impractical Our System: Cheap, Easy to handle for non- biologists in the fjeld, plug-&-play, quantifjable 26

Microbial Biosensors Now: Numerous but Impractical Our System: Cheap, Easy to handle for non- biologists in the fjeld, plug-&-play, quantifjable “Alternative read-outs are very interesting. For a device like yours, which is able to be applied in the fjeld, this is a really nice trick.” - Dr Van Der Burg (BioDetection Systems) 27

B t By ofgering a synbio solution to solve an every day problem, we prove to the broader public that they need synthetic biology in their life. 28

By ofgering a synbio solution to solve an every day problem, we prove to the proader public that they need synthetic biology in their life. “If your system is capable of detecting land mines, I wouldn’t mind if it uses synthetic biology.” - Suzanne Oosterwijk, PAX 29

μtrace - a Custom-designed Device Inexpensive Prototyping 33 μL of ~$3 Analyte Glass Slide Fluid Channel Commercial Electrode Electrical Contacts PDMS 30

μtrace - a Custom-designed Device Inexpensive Prototyping 31

Experimental Setup Outfmow μtrace Counter Electrode Working Electrode Infmow Syringe Reference Electrode Pump Potentiostat A 32

μ trace test results - Ascorbic Acid Voltammetry ybiJ Promoter Ascorbic Acid 0.4V 1.8 Ascorbic Acid 0.2V 1.6 Water 0.4V 1.4 Water 0.2V 1.2 Current ( mA) 1.0 0.8 0.6 0.4 0.2 0 50 100 150 200 250 300 Time (s) 33

μtrace with MtrCAB & Ccm E. coli C43 + MtrCAB & Ccm 0.045 E. coli C43 0.04 Water 0.035 0.03 Current ( μ A) 0.025 0.02 0.015 0.01 0.005 0 50 100 150 200 250 300 Time (s) 34

Achievements Improved responsiveness of modifjed E. coli to detect landmines Stochastic modeling of Curli shows a qualitative response Improved the formerly registered MtrCAB BioBrick Modifjed E. coli to produce an electrical output Designed μtrace - a microfmuidic device for measurement of electrical output 35

e ectrace A plug-&-play platform for biosensors, that produces a quantitative electrical output that can be used for the detection of landmines. 36

Partner: Sponsors: TU Delft Department of Biotechnology TU Delft Faculty of Applied Sciences 37

e ectrace A plug-&-play platform for biosensors, that produces a quantitative electrical output that can be used for the detection of landmines. 38