long-term monitoring of bacteria undergoing programmed population control in a microchemostat
DESCRIPTION
Long-Term Monitoring of Bacteria Undergoing Programmed Population Control in a Microchemostat. Frederick K. Balagaddé, Lingchong You, Carl L. Hansen, Frances H. Arnold, Stephen R. Quake. Science , 309:137-140 (2005). Article critique presented by Véronique Lecault - PowerPoint PPT PresentationTRANSCRIPT
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Long-Term Monitoring of Bacteria Undergoing Programmed Population Control in a Microchemostat
Frederick K. Balagaddé, Lingchong You, Carl L. Hansen, Frances H. Arnold, Stephen R. Quake
Article critique presented by Véronique Lecault
January 10, 2006 EECE 491c
Science, 309:137-140 (2005)
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EECE 491c Balagaddé et al. (2005) Science 309:137-140 2
Microfluidics: A Small World with Large Applications…
Protein crystallization
DNA amplification
Electrophoresis
Immunoassays
DNA microarrays
Cell sorting and analysis
(Thorsen et al., 2002)
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EECE 491c Balagaddé et al. (2005) Science 309:137-140 3
Scaling Bioreactors Down to Nanoliters…
11x10-31x10-61x10-9 1x103 1x106 1x109 Liters
Advantages:
• Reduced reagent costs
• High-throughput experiments
• Cell tracking
• Reduced waste
• Increased speed of experiments
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EECE 491c Balagaddé et al. (2005) Science 309:137-140 4
How do we culture cells?• Batch
• Fed-batch (semi continuous)
• Chemostat (continuous)
Time
Num
ber
of b
acte
ria
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EECE 491c Balagaddé et al. (2005) Science 309:137-140 5
Bacteria and biofilms
(www.medicdirect.co.uk)
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EECE 491c Balagaddé et al. (2005) Science 309:137-140 6
Bacteria and biofilms
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EECE 491c Balagaddé et al. (2005) Science 309:137-140 7
The Microchemostat
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EECE 491c Balagaddé et al. (2005) Science 309:137-140 8
The Microchemostat
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EECE 491c Balagaddé et al. (2005) Science 309:137-140 9
The MicrochemostatExperiment Medium [ glucose]
(M)[bacto-
tryptone] (g/L)
Temperature (oC)
Dilution rate (h-1)
1 MOPS EZ 1.1 -- 32 0.34
2 MOPS EZ 0.11 -- 32 0.30
3 LB -- 0.5 21 0.24
4 LB -- 0.5 21 0.30
5 LB -- 3.0 21 0.37
6 LB -- 0.5 21 0.37
7 LB -- 0.1 21 0.37
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EECE 491c Balagaddé et al. (2005) Science 309:137-140 10
The Microchemostat
3 g/L
0.5 g/L
0.1 g/L
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EECE 491c Balagaddé et al. (2005) Science 309:137-140 11
Quorum Sensing
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EECE 491c Balagaddé et al. (2005) Science 309:137-140 12
Quorum Sensing
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EECE 491c Balagaddé et al. (2005) Science 309:137-140 13
Quorum Sensing
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EECE 491c Balagaddé et al. (2005) Science 309:137-140 14
Quorum Sensing
IPTG -
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EECE 491c Balagaddé et al. (2005) Science 309:137-140 15
Quorum Sensing
IPTG +
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EECE 491c Balagaddé et al. (2005) Science 309:137-140 16
MC4100Z1 Cells
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EECE 491c Balagaddé et al. (2005) Science 309:137-140 17
Top10F’ Cells
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EECE 491c Balagaddé et al. (2005) Science 309:137-140 18
Loss of Regulation
MC4100Z1
Top10F’
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EECE 491c Balagaddé et al. (2005) Science 309:137-140 19
Population dynamics
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EECE 491c Balagaddé et al. (2005) Science 309:137-140 20
Population dynamics
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EECE 491c Balagaddé et al. (2005) Science 309:137-140 21
Conclusions
• Operation of a miniaturized bioreactor 300 times smaller than the previous microfermentor.
• Automatic culturing and monitoring of populations of 100 to 10000 bacteria with instantaneous cell resolution.
• In theory, the mutation rate should be reduced by a factor of 105 and
prolong monitoring of homogeneous populations.
• Monitoring the programmed behavior of bacterial population for hundred of hours despite strong selection pressure.
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EECE 491c Balagaddé et al. (2005) Science 309:137-140 22
Critique Summary
To be improved Good
Ma
jor
po
ints
Min
or
po
ints
•Experiments missing to explain what causes the loss of regulation
•Could fit the model to data in order to estimate the true constants
•Could put more emphasis on the absence of biofilm formation and on the limitations of the chemostat
•Should compare mutation rates to a macroscale chemostat instead of a microscale batch
•Very novel technology
•Attempt to show a possible application
•Honesty of the authors
•Clear representation of the microchemostat technology
•Some figures are not very intuitive to read
•Explanation missing for the morphological behavior
•Incomplete Material and methods section leading to hardly reproducible experiments