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Team:Chiba/Project
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Our project uses two classes of bacteria: senders and receivers.Senders produce signaling molecules, and receivers are activated only after a particular concentration of this molecule is reached.The communication using non-endogenous molecules is less sensitive,and it requires higher signal concentration to take effect.This results in slower activation of receivers. | Our project uses two classes of bacteria: senders and receivers.Senders produce signaling molecules, and receivers are activated only after a particular concentration of this molecule is reached.The communication using non-endogenous molecules is less sensitive,and it requires higher signal concentration to take effect.This results in slower activation of receivers. | ||
| - | *Quorum-Sensing Cross-talk | + | *Sender Phase |
| + | **Quorum-Sensing Cross-talk | ||
AHLs produced by different bacteria differ only in the length of the acyl-chain moiety and substitution at position C-3.([http://partsregistry.org/Part:BBa_F2620:Specificity BBa_F2620:Specificity]) | AHLs produced by different bacteria differ only in the length of the acyl-chain moiety and substitution at position C-3.([http://partsregistry.org/Part:BBa_F2620:Specificity BBa_F2620:Specificity]) | ||
| - | *LuxR/Plux mutants show | + | *Receiver Phase |
| + | **LuxR/Plux mutants show | ||
#a greater response to 3OC6HSL ([http://authors.library.caltech.edu/5553/ C. H. Collins.''et al''.Mol.Microbiol.2005.]) | #a greater response to 3OC6HSL ([http://authors.library.caltech.edu/5553/ C. H. Collins.''et al''.Mol.Microbiol.2005.]) | ||
#a increase in sensitivity to 3OC12HSL ([http://mic.sgmjournals.org/cgi/content/abstract/151/11/3589 B. Koch.''et al''.Microbiology (2005)]). | #a increase in sensitivity to 3OC12HSL ([http://mic.sgmjournals.org/cgi/content/abstract/151/11/3589 B. Koch.''et al''.Microbiology (2005)]). | ||
| - | + | **AHL reporter with aiiA | |
| + | Express LuxR and aiiA constantly. AiiA degrades AHL as signaling molecule. Express GFP when the AHL concentration exceed the capacity of aiiA. | ||
| + | This enables the delay of the activation time of receiver. | ||
==== About Quorum Sensing ==== | ==== About Quorum Sensing ==== | ||
Quorum sensing is a cell-to-cell signaling action of bacteria. They detect the cell density of the same species and coordinate the expression behavior of their cells. Species of Gram-Negative signaling transfer molecules (so-called autoinducer) is a series of acyl homoserine lactone (AHL). The signals are synthesized from S-adenosylmethionine(SAM) by a synthase protein and once they have reached a threshold concentration,they bound to a transcriptional regulatory protein to induce expression of target genes. | Quorum sensing is a cell-to-cell signaling action of bacteria. They detect the cell density of the same species and coordinate the expression behavior of their cells. Species of Gram-Negative signaling transfer molecules (so-called autoinducer) is a series of acyl homoserine lactone (AHL). The signals are synthesized from S-adenosylmethionine(SAM) by a synthase protein and once they have reached a threshold concentration,they bound to a transcriptional regulatory protein to induce expression of target genes. | ||
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==What our system requires== | ==What our system requires== | ||
*Communication module | *Communication module | ||
Revision as of 11:44, 24 October 2008
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Contents |
Introduction
- "Team : Chiba - E.coli time manager"
We control the timing of gene expression by using multiple signaling devices.To this end,we utilize molecules associated with Quorum sensing, a phenomenon that allows bacteria to communicate with each other.Our project uses two classes of bacteria: senders and receivers. Senders produce signaling molecules, and Receivers are activated only after a particular concentration of this molecule is reached.Although different quorum sensing species have slightly different signaling molecules, these molecules are not completely specific to their hosts and cross-species reactivity is observed (M.K Winson et al.FEMS Microbiology Letters,1998). Communication using non-endogenous molecules is less sensitive, and requires a higher signal concentration to take effect.This results in slower activation of receivers.
Motivation
Project Design
Controlling the time of a cell-to-cell signaling action
Our project uses two classes of bacteria: senders and receivers.Senders produce signaling molecules, and receivers are activated only after a particular concentration of this molecule is reached.The communication using non-endogenous molecules is less sensitive,and it requires higher signal concentration to take effect.This results in slower activation of receivers.
- Sender Phase
- Quorum-Sensing Cross-talk
AHLs produced by different bacteria differ only in the length of the acyl-chain moiety and substitution at position C-3.(BBa_F2620:Specificity)
- Receiver Phase
- LuxR/Plux mutants show
- a greater response to 3OC6HSL (C. H. Collins.et al.Mol.Microbiol.2005.)
- a increase in sensitivity to 3OC12HSL (B. Koch.et al.Microbiology (2005)).
- AHL reporter with aiiA
Express LuxR and aiiA constantly. AiiA degrades AHL as signaling molecule. Express GFP when the AHL concentration exceed the capacity of aiiA. This enables the delay of the activation time of receiver.
About Quorum Sensing
Quorum sensing is a cell-to-cell signaling action of bacteria. They detect the cell density of the same species and coordinate the expression behavior of their cells. Species of Gram-Negative signaling transfer molecules (so-called autoinducer) is a series of acyl homoserine lactone (AHL). The signals are synthesized from S-adenosylmethionine(SAM) by a synthase protein and once they have reached a threshold concentration,they bound to a transcriptional regulatory protein to induce expression of target genes.
More about Quorum Sensing
What our system requires
- Communication module
- Quick output
How Our System Works
Experiments
Quorum-Sensing Cross-talk
- Senders
Method
- Transform Senders into E.coli strains(JW1908/XL10GOLD) and Receiver into E.coli strain(JW1908).
- Inoculated them independently in liquid media. Incubated at 37℃ 12h.
- Washed Senders and receiver.
- Mix them.
- Incubated at 37℃ or 30℃.
- Measured intensity of green fluorescence at regular time intervals.
Result
LuxR mutation
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