eau d’e coli igem 2006: mit2006.igem.org/wiki/images/c/c0/igem2006-mit-powerpoint.pdf · 2006....
TRANSCRIPT
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eau d’e coliigem 2006: mit
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meet the teamThe Undergrads:Kate Broadbent Biological Engineering
Andre Green Biology / Physics
Stephen Payne Biological Engineering
Veena Venkatachalam Chemistry / Physics
Boyuan Zhu Electrical Engineering and Computer Science
The Grad Students:Barry CantonAustin CheJason KellyReshma ShettySamantha Sutton
The Faculty:Drew EndyTom Knight
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motivations
eau d’e coli
mit igem 2006
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background
A. thaliana
JMTjasmine
O. basilicum
CCMTcinnamon
A. majus
BAMTfresh / floral
BSMTwintergreen
C. breweri S. cerevisiae
ATF1banana
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background
O. basilicum
CCMTcinnamon
A. majus
BAMTfresh / floral
BSMTwintergreen
C. breweri S. cerevisiae
ATF1banana
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background
A. majus
BAMTfresh / floral
BSMTwintergreen
C. breweri S. cerevisiae
ATF1banana
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background
BSMTwintergreen
C. breweri S. cerevisiae
ATF1banana
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timeline:initial planninginitial planning • scent production • self-generating scent • production regulation • eliminating indole
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timeline:scent productioninitial planning • scent production • self-generating scent • production regulation • eliminating indole
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timeline:self-generating scent
initial planning • scent production • self-generating scent • production regulation • eliminating indole
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timeline:production regulation
initial planning • scent production • self-generating scent • production regulation • eliminating indole
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timeline:indole eliminationinitial planning • scent production • self-generating scent • production regulation • eliminating indole
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wintergreen generating device
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device generates wintergreen
O
O
OH
methyl salicylateMW = 152no methyl salicylate
[no peak at 9.1 minutes]
E. coli E. coli + precursor (SA) + WGD
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banana generating device
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precursor biosynthesis
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precursor biosynthesis
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goal: the regulated system
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growth dependent regulation
osmY: active in stationary phase & under high osmotic pressure conditions
osmY osmY + inverter
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regulation is growth dependent
0
20000
40000
60000
80000
100000
120000
140000
160000
180000
200000
-0.000000007450.099999994040.199999995530.299999997020.399999998510.500000000000.600000001490.70000000298
BBa_J45995 (pOsmY-GFP)BBa_J45017 (Negative control)BBa_I7100 (Constitutive promoter-GFP)
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regulation is growth dependent
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regulated system: exponential
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regulated system: stationary
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regulated system
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chassis (tnaA5-) does not produce indole
No indole [no peak at 10 minutes]
IndolePeak at 10 minutes
MW = 117
wild type E. coli chassis: tnaA5- E. coli
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final system
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smell test results
0
10
20
30
40
50
60
70
80
90
100
A (Mint) B (Stinky) C (Banana)
BananaMintStinky
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future applications
Improve the workplace environment for microbiologists working with E. coli
Port our system to bacterial species involved in bioremediation.
Implement our system in bacteria responsible for bad human odor in the mouth, armpits and feet.
Implement our system in yeast used in bread and beer production
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conclusion
Designed, built, and tested a synthetic biological system over the course of summer
Engineered synthetic scent systems in E. coli
Regulated via transcription control devices
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acknowledgmentsNatalia Dudareva, Purdue University: gifts of expression vectors encoding BAMT, SAMT and BSMT enzymes.
Eran Pichersky, University of Michigan: suggestion of eliminating indole from Escherichia coli to mitigate the natural "bad" smell.
Mary Berlyn, CGSC, The Coli Genetic Stock Center: Esherichia coli strain YYC912.
Brian Cook, MIT: valuable discussions.
Cornelia Reimmann, University of Lausanne: gift of an expression vector carrying the pchBA coding region.
Dieter Haas, University of Lausanne: gift of the Pseudomonas fluorescens strain CHA0.
Peter Bakker, Utrecht University, The Netherlands: gift of an expression vector carrying the pmsCEAB coding region, as well as, gift of the Pseudomonas fluorescens strain WCS374.
Herbert P. Schweizer, Colorado State University: gift of the pUCP22 Esherichia coli to Pseudomonas shuttle vector.
Pamela Silver, Harvard Medical School: gift of yeast BioBricks vector.
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questions and answers
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growth curves
0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0 20,000 40,000 60,000 80,000
Bba_J45995 (pOsmY-GFP)Bba_J45017 (Negative control)BBa_I7100 (Constitutive promoter-GFP)