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![Page 1: “Engineering modular and orthogonal genetic logic gates for robust digital-like synthetic biology” Baojun Wang, Richard I Kitney, Nicolas Joly & Martin.](https://reader031.fdocuments.us/reader031/viewer/2022032309/56649d195503460f949ee908/html5/thumbnails/1.jpg)
“Engineering modular and orthogonal genetic logic gates for
robust digital-like synthetic biology”
Baojun Wang, Richard I Kitney, Nicolas Joly & Martin BuckJamal Elkhader & Queenie Chan
22 February 2012
![Page 2: “Engineering modular and orthogonal genetic logic gates for robust digital-like synthetic biology” Baojun Wang, Richard I Kitney, Nicolas Joly & Martin.](https://reader031.fdocuments.us/reader031/viewer/2022032309/56649d195503460f949ee908/html5/thumbnails/2.jpg)
Background
AND gate:
![Page 3: “Engineering modular and orthogonal genetic logic gates for robust digital-like synthetic biology” Baojun Wang, Richard I Kitney, Nicolas Joly & Martin.](https://reader031.fdocuments.us/reader031/viewer/2022032309/56649d195503460f949ee908/html5/thumbnails/3.jpg)
Background
NAND gate:
![Page 4: “Engineering modular and orthogonal genetic logic gates for robust digital-like synthetic biology” Baojun Wang, Richard I Kitney, Nicolas Joly & Martin.](https://reader031.fdocuments.us/reader031/viewer/2022032309/56649d195503460f949ee908/html5/thumbnails/4.jpg)
Background
Terms:o Digital system: on/off
o Forward engineering: classifying/evaluating components before choosing the parts to build the system
o Modular: can change inputs and outputs and logic gate will still work
o Orthogonal: no cross-talk
![Page 5: “Engineering modular and orthogonal genetic logic gates for robust digital-like synthetic biology” Baojun Wang, Richard I Kitney, Nicolas Joly & Martin.](https://reader031.fdocuments.us/reader031/viewer/2022032309/56649d195503460f949ee908/html5/thumbnails/5.jpg)
Project Overview
Purpose: to engineer a digital-like system using modular and orthogonal parts via forward engineering
I. ClassificationII. Design AND gateIII.Test ModularityIV. repeat II and III for NAND gateV. Compatibility
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Classification: Forward Engineering
• 3 different promoters: Plac, PBAD, Plux
• 3 test conditions:
1. promoters' responses, varying chassis (E. coli MC4100, E. coli MC1061)
IPTG --> lacArabinose --> BADAHL --> lux
Classification Design AND gate
Test Modularity NAND gate Compatibilit
y
![Page 7: “Engineering modular and orthogonal genetic logic gates for robust digital-like synthetic biology” Baojun Wang, Richard I Kitney, Nicolas Joly & Martin.](https://reader031.fdocuments.us/reader031/viewer/2022032309/56649d195503460f949ee908/html5/thumbnails/7.jpg)
Classification: Forward Engineering
• 3 different promoters: Plac, PBAD, Plux
• 3 test conditions:
1. promoters' responses, varying chassis (E. coli MC4100, E. coli MC1061)
2. promoters + RBSs responses with varying RBS's
IPTG --> lacArabinose --> BADAHL --> lux
Classification Design AND gate
Test Modularity NAND gate Compatibilit
y
![Page 8: “Engineering modular and orthogonal genetic logic gates for robust digital-like synthetic biology” Baojun Wang, Richard I Kitney, Nicolas Joly & Martin.](https://reader031.fdocuments.us/reader031/viewer/2022032309/56649d195503460f949ee908/html5/thumbnails/8.jpg)
Classification: Forward Engineering
• 3 different promoters: Plac, PBAD, Plux
• 3 test conditions:
1. promoters' responses, varying chassis (E. coli MC4100, E. coli MC1061)
2. promoters + RBSs responses with varying RBS's
3. Promoters + RBS's responses with varying temperatures
IPTG --> lacArabinose --> BADAHL --> lux
Classification Design AND gate
Test Modularity NAND gate Compatibilit
y
![Page 9: “Engineering modular and orthogonal genetic logic gates for robust digital-like synthetic biology” Baojun Wang, Richard I Kitney, Nicolas Joly & Martin.](https://reader031.fdocuments.us/reader031/viewer/2022032309/56649d195503460f949ee908/html5/thumbnails/9.jpg)
Circuit Design: AND Gates
Gate 1
Gate 2
Testing for Modularity
Classification Design AND gate
Test Modularity NAND gate Compatibilit
y
![Page 10: “Engineering modular and orthogonal genetic logic gates for robust digital-like synthetic biology” Baojun Wang, Richard I Kitney, Nicolas Joly & Martin.](https://reader031.fdocuments.us/reader031/viewer/2022032309/56649d195503460f949ee908/html5/thumbnails/10.jpg)
Gate 2
30ºC
37ºC
![Page 11: “Engineering modular and orthogonal genetic logic gates for robust digital-like synthetic biology” Baojun Wang, Richard I Kitney, Nicolas Joly & Martin.](https://reader031.fdocuments.us/reader031/viewer/2022032309/56649d195503460f949ee908/html5/thumbnails/11.jpg)
NAND gates: Design & Modularity Test
Gate 3
Gate 4
Classification Design AND gate
Test Modularity NAND gate Compatibilit
y
![Page 12: “Engineering modular and orthogonal genetic logic gates for robust digital-like synthetic biology” Baojun Wang, Richard I Kitney, Nicolas Joly & Martin.](https://reader031.fdocuments.us/reader031/viewer/2022032309/56649d195503460f949ee908/html5/thumbnails/12.jpg)
Gate 3 Gate 4
Predicted
Results
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NAND gate FACS data
Both inputs added -->
off
Classification Design AND gate
Test Modularity NAND gate Compatibilit
y
![Page 14: “Engineering modular and orthogonal genetic logic gates for robust digital-like synthetic biology” Baojun Wang, Richard I Kitney, Nicolas Joly & Martin.](https://reader031.fdocuments.us/reader031/viewer/2022032309/56649d195503460f949ee908/html5/thumbnails/14.jpg)
• Compatibility of AND gate with chassis
• Top: Gate 1 (Plac + PBAD)
• Bottom: Gate 2 (Plux + PBAD)
Compatibility✗
✗ ✗✗
✗
✗
Classification Design AND gate
Test Modularity NAND gate Compatibilit
y
![Page 15: “Engineering modular and orthogonal genetic logic gates for robust digital-like synthetic biology” Baojun Wang, Richard I Kitney, Nicolas Joly & Martin.](https://reader031.fdocuments.us/reader031/viewer/2022032309/56649d195503460f949ee908/html5/thumbnails/15.jpg)
Conclusions
• AND and NAND gates are modular
• Sigmoidal response property allows system to be robust (minimal background noise)
• Forward engineering and engineering-by-parts works!
• Crosstalk can be avoided by picking promoters that are not native to chassis
![Page 16: “Engineering modular and orthogonal genetic logic gates for robust digital-like synthetic biology” Baojun Wang, Richard I Kitney, Nicolas Joly & Martin.](https://reader031.fdocuments.us/reader031/viewer/2022032309/56649d195503460f949ee908/html5/thumbnails/16.jpg)
SignificanceModular:
• Can use other inputs and outputs and AND / NAND gate will still work
• Parts-based engineering approach
Control:
• New genetic logic devices can be engineered predictably to generate desired behaviours in response to specific extra- or intra- cellular signalling inputs
• Improves sensing specificity and accuracy of biological control
![Page 17: “Engineering modular and orthogonal genetic logic gates for robust digital-like synthetic biology” Baojun Wang, Richard I Kitney, Nicolas Joly & Martin.](https://reader031.fdocuments.us/reader031/viewer/2022032309/56649d195503460f949ee908/html5/thumbnails/17.jpg)
Supplementary Slides
PlacPBAD
Plux