Team:Tokyo Tech/Project/wolf coli

From 2010.igem.org

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(Wolf coli. and Artificial Cooperative System)
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<td>[[Team:Tokyo_Tech|1 Graphical abstract]]<br>
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<td>[[Team:Tokyo_Tech|1 Graphic abstract]]<br>
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<th>4 wolfcoli overview  -YOU ARE HERE!-<br>
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<th>4 Wolf coli overview  -YOU ARE HERE!-<br>
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:[[Team:Tokyo_Tech/Project/wolf_coli/New_Series_of_PompC|4-1 the new seriesof P''ompC'']]
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:[[Team:Tokyo_Tech/Project/wolf_coli/New_Series_of_PompC|4-1 New seriesof P''ompC'']]
:[[Team:Tokyo_Tech/Project/wolf_coli/lacIM1|4-2 lacIM1 for band-detect network ]]
:[[Team:Tokyo_Tech/Project/wolf_coli/lacIM1|4-2 lacIM1 for band-detect network ]]
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:[[Team:Tokyo_Tech/Project/wolf_coli/System|4-3 wolfcoli system]]
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:[[Team:Tokyo_Tech/Project/wolf_coli/System|4-3 Wolf coli system]]
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<font size="5"><b>4 Wolf coli. Overview</b></font>
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<font size="5"><b>4 Wolf coli Overview</b></font>
==Introduction==
==Introduction==
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Have you heard the legend of 'The Wolfman'? They're ordinary man at daytime, but suddenly transform into a ferocious wolf in the full-moon night. Our project aim to imitate the character of Wolfman, more specifically, designing two types of E.coli that helps each other to survive at daytime, whereas competing at full moon night.
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[[Image:tokyotech_wolfcoli_system_ver2.png|left|thumb|250px|Fig. 4-1 Cooperative activity of Artificial Cooperation System]]
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[[Image:tokyotech_wolfcoli_system_ver5.png|right|thumb|350px|Figure 4-2. Overview of “Wolf coli” system]]
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[[Image:Tokyotech_wolfcoli_ver3.png|left|thumb|250px|Figure3-1. Introduction of “Wolf coli.”]]
 
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[[Image:Tokyotech wolfcoli system.png|left|thumb|300px|Figure3-2. Overview of “Wolfcoli” system]]
 
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Have you heard the legend of 'The Wolfman'? They're ordinary man at daytime, but suddenly transform into a ferocious wolf in the full-moon night. Our project aim to imitate the character of Wolfman, more specifically, designing two types of ''E.coli'' that helps each other to survive at daytime, whereas competing at full moon night. In order to create the “Wolf coli”, we introduced " red-light-dependent gene expression network"[1] and "band-detect network"[2] into one cell, and combined these networks with the Artificial Cooperation System (Fig. 4-3). We characterized new series of ''OmpC'' promoter, and LacIM1 which are crucial parts of  our networks.
 +
[[Image:Tokyotech_ompc_graph.jpg|thumb|left|320px|Fig. 4-3 The induction of new P''ompC'' series in high osmolarity medium at 4 hours. This work is done by Thiprampai THAMAMONGOOD and Taichi NAKAMURA]]
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[[Image:Tokyotech_LacIM1_data.png|thumb|right|230px|Fig. 4-4 Repression efficiency of LacIM1 (BBa_K395401) / LacIWT (BBa_K395402) exposed to arabinose and IPTG.  This work is done by Mitsuhiko ODERA]]
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In order to create the “Wolf coli.”, we introduced " red-light-dependent gene expression network"[1] and "band-detect network"[2] and combined these networks with the Artificial Cooperation System (Fig. 3-2). We conducted experiment on series of OmpC promoter (PompC), and LacIM1 which are crucial parts of  our networks.
 
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[[Image:Tokyotech_ompc_graph.jpg|thumb|left|320px|Figure3-3. The induction of new P''ompC'' series in high osmolarity medium at hours. This work is done by Thiprampai THAMAMONGOOD and Taichi NAKAMURA]]
 
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[[Image:Tokyotech_LacIM1_data.png|thumb|right|230px|Figure3-4.Repression efficiency of LacIM1 (BBa_K395401) / LacIWT (BBa_K395402) exposed to arabinose and IPTG.  This work is done by Mitsuhiko ODERA]]
 
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We have succeeded in designing 2 new osmoregulative promoters, P''ompC(CB)'' [http://partsregistry.org/Part:BBa_K395302 (BBa_K395302)] and P''ompC(CS1)''[http://partsregistry.org/Part:BBa_K395303 (BBa_K395303)], which can be utilized in the red-light-dependent gene expression network (Fig. 4-3). We also characterized LacIM1 [http://partsregistry.org/Part:BBa_K082026 (BBa_K082026)], a mutant of LacIWT,which is a key component in the band-detect network. Although, this part was registered by USTC(2008) [3], it was not well characterized in the BioBrick registry. We confirmed that LacIM1 shows weaker repression to Plac than its wild type. (Fig. 4-4)
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==Wolf coli and Artificial Cooperative System ==
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[[Image:tokyotech_wolfcoli_system_ver2.png|thumb|center|350px|Fig. 4-1 Cooperative activity of Artificial Cooperation System]]
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The Artificial Cooperation System was designed to be switched off during the “full moon night”. Therefore, Sympathetic coli would  transform into Wolf coli at the “full moon light”. During this period, communication between 2 types of cells doesn't occur. Hence, both types of cells are unable to recognize each other by quorum sensing and become competitors. Cooperative activity in Artificial Cooperation System is regulated by 3 levels of light intensity which are weak, medium and strong. “Weak light” from the crescent moon switches the Artificial Cooperation System on, thus two types of cells are able to communicate and help each other. “Medium light” during the full moon night can switch the system off resulting in appearance of the “Wolfcoli”. During the daytime, “strong light” from the sun activates the Artificial Cooperation System.
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==P''OmpC'' in red-light-dependent gene expression network==
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[[Image:Tokyotech wolfcoli system_ver5.png|left|thumb|300px|Fig. 4-5 Overview of red-light-dependent gene expression network in Wolf coli system]]
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[[Image:Tokyotech_ompc_graph.jpg|thumb|left|320px|Fig. 4-6 The activation of new P''ompC'' series in high osmolarity medium at 4 hours. This work is done by Thiprampai THAMAMONGOOD and Taichi NAKAMURA]]
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''OmpC'' promoter, in red-light-dependent gene expression network, plays a crucial role in initiating the transformation of Sympathetic coli into Wolf coli. To accomplish band-detect circuit in light sensing system, varieties of ''OmpC'' promoters were designed and characterized so as to find an appropriate strength of the promoter particularly which can be activated by the “full moon light”.(Fig.4-5)
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[http://2010.igem.org/Team:Tokyo_Tech/Project/wolf_coli/New_Series_of_PompC#Introduction ...see more about ''OmpC'' promoter ](Fig. 4-6)
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==LacIMI in band-detect network==
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[[image:Tokyotech_band_detect.png|340px|thumb|left| Fig.4-7 Patial circuit of band-detect network ]]
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[[Image:Tokyotech_LacIM1_data.png|thumb|right|240px|Fig. 4-8.Repression efficiency of LacIM1 (BBa_K395401) / LacIWT (BBa_K395402) exposed to arabinose and IPTG.  This work is done by Mitsuhiko ODERA]]
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We succeeded in designing 2 new osmoregulative promoters, PompC(CB) (BBa_K395302) and PompC(CS1) (BBa_K395303), which can be utilized in the red-light-dependent gene expression network (Fig. 3-3).
 
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We characterized lacIM1 (BBa_K082026), a mutation of LacI (BBa_?????),which is a key component in the band-detect network. In fact, this part was registered by USTC(2008) [3], however it was not well characterized in the BioBrick registry. We found that product of lacIM1 shows weaker repression to Plac than its wild type. (Fig. 3-4)
 
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==Wolf coli. and Artificial Cooperative System ==
 
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[[image:Tokyotech_Cooperative_activity.jpg|center|300px]]
 
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The Aftificial Cooperation System was designed to be switching off during the “full moon night” which means sympathetic coli. would be transformed into wolf coli. by induction of the “full moon light”. During this period, communication between 2 types of cells is inhibited. Therefore, both types of cells are competitors and they are unable to recognize each other by quorum sensing. Cooperative activity in Artificial Cooperation System is regulated by 3 levels of light intensity which are weak, medium and strong. “Weak light” from the crescent moon enables the Artificial Cooperation System on, thus two types of cell are able to communicate and help each other. “Medium light” during the full moon night can switch the system off resulting in declining of the cooperative activity and transforming of sympathetic coli. into “Wolf coli”. During the daytime, “strong light” from the sun switches the system on then quorum sensing is activated.
 
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==P''OmpC'' in red-light-dependent gene expression network==
 
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[[Image:Tokyotech wolfcoli system.png|300px|Figure3-2. Overview of “Wolfcoli” system]]
 
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[[image:Tokyotech_Band_detector_circuit.JPG|300px|PompC(WT) and the two-component system]]
 
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[http://2010.igem.org/Team:Tokyo_Tech/Project/wolf_coli/New_Series_of_PompC#Introduction ''OmpC'' promoter], in red-light-dependent gene expression network, plays a crucial role in initiating the transformation of sympathetic coli. into Werewolf coli. in light sensitive system. To accomplish band-detect circuit in light sensitive system, varieties of OmpC promoters were designed and characterized in order to find an appropriate strength of the promoter particularly which can be activated by the “full moon light”.
 
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==LacIMI in band-detect network==
 
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[[Image:Tokyotech wolfcoli system.png|left|thumb|300px|Figure3-2. Overview of “Wolfcoli” system]]
 
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The band-detect network exhibits transient gene expression in response to concentration of chemical signals(Fig.4-7). In the band-detect network, LacIM1 is a crucial component due to its low repression efficiency[1]. According to the assay results, we confirmed that LacIM1 shows weaker repression to Plac than its wild type.(Fig. 4-8). [http://2010.igem.org/Team:Tokyo_Tech/Project/wolf_coli/lacIM1 …see more about lacIM1].
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==References==
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[1]. Basu S, Gerchman Y, Collins CH, et al. A synthetic multicellular system for programmed pattern formation. NATURE 2005;434,1130-1134
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[http://2010.igem.org/Team:Tokyo_Tech/Project/wolf_coli/lacIM1 LacIM1] is a crucial component in the band-detect network (Fig. 3-2).
 
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Latest revision as of 03:56, 28 October 2010

iGEM Tokyo Tech 2010 "E.coli with Humanity"

4 Wolf coli Overview

Contents

Introduction

Fig. 4-1 Cooperative activity of Artificial Cooperation System
Figure 4-2. Overview of “Wolf coli” system












Have you heard the legend of 'The Wolfman'? They're ordinary man at daytime, but suddenly transform into a ferocious wolf in the full-moon night. Our project aim to imitate the character of Wolfman, more specifically, designing two types of E.coli that helps each other to survive at daytime, whereas competing at full moon night. In order to create the “Wolf coli”, we introduced " red-light-dependent gene expression network"[1] and "band-detect network"[2] into one cell, and combined these networks with the Artificial Cooperation System (Fig. 4-3). We characterized new series of OmpC promoter, and LacIM1 which are crucial parts of our networks.

Fig. 4-3 The induction of new PompC series in high osmolarity medium at 4 hours. This work is done by Thiprampai THAMAMONGOOD and Taichi NAKAMURA
Fig. 4-4 Repression efficiency of LacIM1 (BBa_K395401) / LacIWT (BBa_K395402) exposed to arabinose and IPTG. This work is done by Mitsuhiko ODERA












We have succeeded in designing 2 new osmoregulative promoters, PompC(CB) (BBa_K395302) and PompC(CS1)(BBa_K395303), which can be utilized in the red-light-dependent gene expression network (Fig. 4-3). We also characterized LacIM1 (BBa_K082026), a mutant of LacIWT,which is a key component in the band-detect network. Although, this part was registered by USTC(2008) [3], it was not well characterized in the BioBrick registry. We confirmed that LacIM1 shows weaker repression to Plac than its wild type. (Fig. 4-4)

Wolf coli and Artificial Cooperative System

Fig. 4-1 Cooperative activity of Artificial Cooperation System

The Artificial Cooperation System was designed to be switched off during the “full moon night”. Therefore, Sympathetic coli would transform into Wolf coli at the “full moon light”. During this period, communication between 2 types of cells doesn't occur. Hence, both types of cells are unable to recognize each other by quorum sensing and become competitors. Cooperative activity in Artificial Cooperation System is regulated by 3 levels of light intensity which are weak, medium and strong. “Weak light” from the crescent moon switches the Artificial Cooperation System on, thus two types of cells are able to communicate and help each other. “Medium light” during the full moon night can switch the system off resulting in appearance of the “Wolfcoli”. During the daytime, “strong light” from the sun activates the Artificial Cooperation System.

POmpC in red-light-dependent gene expression network

Fig. 4-5 Overview of red-light-dependent gene expression network in Wolf coli system
Fig. 4-6 The activation of new PompC series in high osmolarity medium at 4 hours. This work is done by Thiprampai THAMAMONGOOD and Taichi NAKAMURA


OmpC promoter, in red-light-dependent gene expression network, plays a crucial role in initiating the transformation of Sympathetic coli into Wolf coli. To accomplish band-detect circuit in light sensing system, varieties of OmpC promoters were designed and characterized so as to find an appropriate strength of the promoter particularly which can be activated by the “full moon light”.(Fig.4-5) ...see more about OmpC promoter (Fig. 4-6)

LacIMI in band-detect network

Fig.4-7 Patial circuit of band-detect network
Fig. 4-8.Repression efficiency of LacIM1 (BBa_K395401) / LacIWT (BBa_K395402) exposed to arabinose and IPTG. This work is done by Mitsuhiko ODERA













The band-detect network exhibits transient gene expression in response to concentration of chemical signals(Fig.4-7). In the band-detect network, LacIM1 is a crucial component due to its low repression efficiency[1]. According to the assay results, we confirmed that LacIM1 shows weaker repression to Plac than its wild type.(Fig. 4-8). …see more about lacIM1.

References

[1]. Basu S, Gerchman Y, Collins CH, et al. A synthetic multicellular system for programmed pattern formation. NATURE 2005;434,1130-1134