Team:Tokyo Tech

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Project overview

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A love story contains several processes. Two people fall in love and their love burning wildly. However, no forever exists in the world, in most occasions, love will eventually burn to only a pile of ashes of the last remaining wind drift away. In our project, we have recreated the story of "Romeo and Juliet" by Shakespeare vividly by two kinds of Escherichia coli cell-cell communication. We aim to generate a circuit involving regulatory mechanism of positive feedback rather than commonly-used negative feedback to control the fate of E.coli by signaling between two types of E.coli. Besides, Rose represents love. We are the first iGEM group ever to synthesize PHAs (a kind of bio-plastics) by using our new biobrick part, representing rose .

Ⅰ cell-cell communication

"Romeo and Juliet" is the dramas by dramatist William Shakespeare of England. The stage is city Verona, Italy in the 14th century. Romeo met Juliet. Two people fell in love instantly. In this project, we will recreate the love story of "Romeo and Juliet".


[Detailed descriptions for Cell-cell communication]

Ⅰ-1 story

We make our cute E.coli play “Romeo and Juliet” which is one of Shakespeare’s most famous plays. In this project, we define the signal that E.coli produce as the romantic feeling of Romeo and Juliet. In this project, we will recreate the love story of "Romeo and Juliet", by using "Cell-cell communication" The story that we reproduce is divided into four scenes.

Click on the Scene.

Scene1

Scene2

Scene3

Scene4

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(Scene 1) Romeo meets and falls in love with Juliet. Once the love between two people stimulates each other, they become deeply attached and cannot live without each other.


First, to represent the condition that their love stimulates each other in “Scene1 Fall in love”, we designed a positive feedback system in which the production of a signal activates the production of the other signal. [Detailed descriptions for Positive feedback system]



Ⅰ-2 Positive Feedback of cell-cell communication

Fig1-2,positive feedback assay

To accomplish the positive feedback system in the cell-cell communication, we designed and constructed two types of genetically engineered E.coli, 3OC6HSL-dependent 3OC12HSL producer cell (Plux-LasI cell) and 3OC12HSL-dependent 3OC6HSL producer cell (Plas-LuxI cell). By co-culturing Plux-LasI cell and Plas-LuxI cell, higher concentration of a signal than that in initial conditions was detected through the production of the other signal (Fig1-2 red arrow & blue arrow). This result strongly suggests that our positive feedback system worked accurately. In the process of the implementation, we also confirmed two modules, the 3OC6HSL-dependent 3OC12HSL production module and the 3OC12HSL-dependent 3OC6HSL production module, which constituted our positive feedback system. In these module, two new Biobrick parts, Plux-LasI ([http://partsregistry.org/wiki/index.php?title=Part:BBa_K934022 BBa_K934022]) and Plas-LuxI ([http://partsregistry.org/wiki/index.php?title=Part:BBa_K934012 BBa_K934012]) , were characterized their functions. [Detailed descriptions for Cell-cell communication]

Other experiments for basis of "Romeo & Juliet"













Ⅰ-3 Modeling

Fig1-6, time-dependent change of the concentrations of the two signals.
Fig1-7: the story of “Romeo and Juliet”

Result: Whether our circuit can reproduce “Romeo and Juliet”

To confirm the feasibility of the cell-cell communication system, we developed an ordinary differential equation model and simulated the system under typical experimental conditions. Fig1-6 shows the result of the simulation. As described below, the behavior of the signal concentration is consistent with the development of the “Romeo and Juliet” story.

In the gray area of Fig1-6, the concentration of two signals increases. It represents Scene1.

In the green area of Fig1-6, as the concentration of Romeo signals increases to the some extent, the concentration of Juliet signals starts to decline. It represents Scene2.

In the blue area of Fig1-6, lysis gene is expressed in Romeo cell in response to the decline of the concentration of Juliet signals, then the concentration of Romeo signals starts to decline. It represents Scene3.

In the pink area of Fig1-6, lysis gene is expressed in Juliet cell in response to the decline of the concentration of Romeo signals, then the concentration of Juliet signals decreases further. It represents Scene4.

[Detailed descriptions for Modeling]



Ⅱ PHAs Production

Ⅱ-1 Story

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There is what of the famous scene of "Romeo and Juliet"

JULIET: O Romeo, Romeo! why are you Romeo? Deny your father and refuse your name;
ROMEO: Shall I hear more, or shall I speak at this?
JULIET: It's but your name that is my enemy; you are yourself, though not a Montague. O, be some other name! What's in a name? that which we call a rose By any other name would smell as sweet.

we will recreate the rose come out in the lines of the famous drama "Romeo and Juliet" by the synthesis of PHAs.

Ⅱ PHAs Production

Ⅱ-1 Story

fig1-8,Rose silhouette on the LB agar plate containing Nile red.

There is what of the famous scene of "Romeo and Juliet"

JULIET: O Romeo, Romeo! why are you Romeo? Deny your father and refuse your name;
ROMEO: Shall I hear more, or shall I speak at this?
JULIET: It's but your name that is my enemy; you are yourself, though not a Montague. O, be some other name! What's in a name? that which we call a rose By any other name would smell as sweet.

we will recreate the rose come out in the lines of the famous drama "Romeo and Juliet" by the synthesis of PHAs.

Ⅱ-2 PHAs production

We made a new biobrick part and succeeded synthesizing Polyhydroixyalkanoates(PHAs). This is the first Biobrick part to synthesize PHAs. In our project, we also draw rose silhouette to produce the balcony scene of “Romeo and Juliet” by the synthesis of PHAs.[Detailed descriptions for PHAs production]

Human Practice

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Taku Nakayama, and Mai Miura (members of Tokyo_Tech iGEM team) have participated in a science cafe as assistants for the event. The two members supported people who are not specialist in biology to plan an imaginary synthetic biology project, to be evaluated from public points of view, and to upgrade the project in accordance with the evaluations.

[Detailed descriptions for Human practice ]