Team:Tokyo Tech

Our project

We want to make a Coli Touch!!

→We try to construct a bacterial ‘touch panel’ which is colored by the pressure.

We name it Coli touch.

Until iGEM2007, input style was only heat, small molecule and light. These input style have problems. For example, Heat input occur thermal gradient, small molecule input occur concentration gradient, light input depend on location of a container. So, we thought input style except these. That is “press”. Press input has an advantage that input signal travel uniformly in a container.

Why pressure?

Example of input style to genetic circuit

- Heat: thermal gradient

- Small molecule: concentration gradient

- Light: strength depend on location of container

        

           Difficulty in uniform introduction

 

  Press!  Confirmatory experiment

Prospect of technological application!!

 

Pressure inducible promoter - P_lac

The affinity of LacI for the lac operator is known to decrease due to a tetramer to dimer transition of LacI

Strategy

Equipment. Instrument for pressure experiment

Fig 4: Fig4-1 is a pressure vessel. Fig4-4 is a pressurizing system. Fig4-2 show a inside of the vessel. The inside is filled with water and the withstand pressure container is put into the inside. Fig4-3 show a water bath placed some vessel.

Strategy 1.Construction

Confirmatory experiment

Fig1:Pressure-response of Plac-GFP under several pressures measured by flow cytometer

  

Fig2-1:Plac-GFP under 30MPa       Fig2-2:Plac-GFP under 0.1MPa       Fig2-3:⊿-GFP under 30MPa

Stratagy 2. Basic device

Stratagy 3. Development of low pressure inducible promoter

We performed random mutagenesis to lac promoter and tried to screen promoters with FACS.

 

 

 

 

 

 

 

Strategy for Isolation of promoters

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Future work

•       try Step 2 and 3

•       characterize isolated promoters

 

4.Genetic toggle switch to implement rewritable function

 

 

 

 

 

 

 

・ genetic toggle switch

       - Writable function

P_lac is induced under 30 MPa pressure

(P_L is not induced by 30 MPa pressure)

Express GFP, CI, and CI represses P_L

→ bright !!

- Erasable function

P_L is induced by heat

Express LacI, P_lac represses GFP

→ go out little by little…

 

Future work

      - Identify pressure model parameter from experimental data

      - Simulate pressure induction by pressure model and estimate

        promoter-RBS strength to implement rewritable function

 

 

 

 

 

 

 

 

 

 

 

 

Mathematical　model