eau d'e coli igem 2006: mit
eau d'e coli igem 2006: mit
eau d'e coli igem 2006: mit
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<strong>eau</strong> d’e <strong>coli</strong><br />
<strong>igem</strong> <strong>2006</strong>: <strong>mit</strong>
meet the team<br />
The Undergrads:<br />
Kate Broadbent<br />
Biological Engineering<br />
Andre Green<br />
Biology / Physics<br />
Stephen Payne<br />
Biological Engineering<br />
Veena Venkatachalam<br />
Chemistry / Physics<br />
Boyuan Zhu<br />
Electrical Engineering and<br />
Computer Science<br />
The Grad Students:<br />
Barry Canton<br />
Austin Che<br />
Jason Kelly<br />
Reshma Shetty<br />
Samantha Sutton<br />
The Faculty:<br />
Drew Endy<br />
Tom Knight
motivations<br />
<strong>eau</strong> d’e <strong>coli</strong><br />
<strong>mit</strong> <strong>igem</strong><br />
<strong>2006</strong>
ackground<br />
A. thaliana<br />
JMT<br />
jasmine<br />
O. basilicum<br />
CCMT<br />
cinnamon<br />
A. majus<br />
BAMT<br />
fresh / floral<br />
C. breweri S. cerevisiae<br />
BSMT<br />
wintergreen<br />
ATF1<br />
banana
ackground<br />
O. basilicum<br />
CCMT<br />
cinnamon<br />
A. majus<br />
BAMT<br />
fresh / floral<br />
C. breweri S. cerevisiae<br />
BSMT<br />
wintergreen<br />
ATF1<br />
banana
ackground<br />
A. majus<br />
BAMT<br />
fresh / floral<br />
C. breweri S. cerevisiae<br />
BSMT<br />
wintergreen<br />
ATF1<br />
banana
ackground<br />
C. breweri S. cerevisiae<br />
BSMT<br />
wintergreen<br />
ATF1<br />
banana
timeline:<br />
initial planning<br />
initial planning • scent production • self-generating scent • production regulation • eliminating indole
timeline:<br />
scent production<br />
initial planning • scent production • self-generating scent • production regulation • eliminating indole
timeline:<br />
self-generating scent<br />
initial planning • scent production • self-generating scent • production regulation • eliminating indole
timeline:<br />
production regulation<br />
initial planning • scent production • self-generating scent • production regulation • eliminating indole
timeline:<br />
indole elimination<br />
initial planning • scent production • self-generating scent • production regulation • eliminating indole
wintergreen generating device
device generates<br />
wintergreen<br />
E. <strong>coli</strong><br />
no methyl salicylate<br />
[no peak at 9.1<br />
minutes]<br />
E. <strong>coli</strong> + precursor (SA)<br />
+ WGD<br />
methyl salicylate<br />
MW = 152<br />
OH<br />
O<br />
O
anana generating device
precursor biosynthesis
precursor biosynthesis
goal: the regulated system
growth dependent regulation<br />
osmY: active in stationary phase & under high osmotic pressure conditions<br />
osmY osmY + inverter
egulation is growth dependent<br />
200000<br />
180000<br />
160000<br />
140000<br />
120000<br />
100000<br />
80000<br />
60000<br />
40000<br />
20000<br />
0<br />
BBa_J45995 (pOsmY-GFP)<br />
BBa_J45017 (Negative control)<br />
BBa_I7100 (Constitutive promoter-GFP)<br />
-0.00000000745 0.09999999404 0.19999999553 0.29999999702 0.39999999851 0.50000000000 0.60000000149 0.70000000298
egulation is growth dependent
egulated system: exponential
egulated system: stationary
egulated system
chassis (tnaA5 - ) does not produce indole<br />
wild type E. <strong>coli</strong> chassis: tnaA5 - E. <strong>coli</strong><br />
Indole<br />
Peak at 10 minutes<br />
MW = 117<br />
No indole<br />
[no peak at 10 minutes]
final system
smell test results<br />
100<br />
90<br />
80<br />
70<br />
60<br />
50<br />
40<br />
30<br />
20<br />
10<br />
0<br />
A (Mint) B (Stinky) C (Banana)<br />
Banana<br />
Mint<br />
Stinky
future applications<br />
Improve the workplace environment for<br />
microbiologists working with E. <strong>coli</strong><br />
Port our system to bacterial species involved in<br />
bioremediation.<br />
Implement our system in bacteria responsible for<br />
bad human odor in the mouth, armpits and feet.<br />
Implement our system in yeast used in bread and<br />
beer production
conclusion<br />
Designed, built, and tested a synthetic biological<br />
system over the course of summer<br />
Engineered synthetic scent systems in E. <strong>coli</strong><br />
Regulated via transcription control devices
acknowledgments<br />
Natalia Dudareva, Purdue University: gifts of expression vectors encoding BAMT, SAMT<br />
and BSMT enzymes.<br />
Eran Pichersky, University of Michigan: suggestion of eliminating indole from Escherichia<br />
<strong>coli</strong> to <strong>mit</strong>igate the natural "bad" smell.<br />
Mary Berlyn, CGSC, The Coli Genetic Stock Center: Esherichia <strong>coli</strong> strain YYC912.<br />
Brian Cook, MIT: valuable discussions.<br />
Cornelia Reimmann, University of Lausanne: gift of an expression vector carrying the<br />
pchBA coding region.<br />
Dieter Haas, University of Lausanne: gift of the Pseudomonas fluorescens strain CHA0.<br />
Peter Bakker, Utrecht University, The Netherlands: gift of an expression vector carrying the<br />
pmsCEAB coding region, as well as, gift of the Pseudomonas fluorescens strain WCS374.<br />
Herbert P. Schweizer, Colorado State University: gift of the pUCP22 Esherichia <strong>coli</strong> to<br />
Pseudomonas shuttle vector.<br />
Pamela Silver, Harvard Medical School: gift of yeast BioBricks vector.
questions and answers
growth curves<br />
0.7<br />
0.6<br />
0.5<br />
0.4<br />
0.3<br />
0.2<br />
0.1<br />
0<br />
Bba_J45995 (pOsmY-GFP)<br />
Bba_J45017 (Negative control)<br />
BBa_I7100 (Constitutive promoter-GFP)<br />
0 20,000 40,000 60,000 80,000
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