Team:Imperial College/Summary

In order to achieve our specifications of design, we require the following devices;

• Light sensing device - Converting a light input into a PoPS output
• Biomaterial production device - Converting a PoPS input into an output of biomaterial production
• Motility Control device - Converting a PoPS input into an output of motility arrest
• Integration device - To allow integration and selection of our genetic constructs and devices into B,subtilis

Each of these constructs makes up the final device which is shown below:

(AB is our antibiotic resistance cassette, ytvA is the gene controlling the light-sensing pathway, SB is the biomaterial, epsE the clutch and the 5' and 3' sections are integration sites. Light-inducible promoters are labelled with an 'L')

Growth Curve

The growth of B. subtilis was characterised by simulating it on MATLAB. The aim was to increase the predictability of its behaviour during the growth. Also, the characterisation acts as a way to verify the accuracy of the experimental results. Several parameters, such as the growth constant and the Hill coefficient, were determined. These were incorporated into the model to further enhance its accuracy.

Genetic Circuit

We have considered two mathematical models describing the time evolution of aspects of the genetic circuitry that comprise our device. We verify which model best describes the behaviour of the circuit using laboratory data.

Motility Analysis

We have developed a simple mechanical model for the swimming motility of B. subtilis. Using manual tracking, we were able to extract x,y coordinate data from the cell trajectory. This allowed us to fit experimental data with our model. It was concluded that flagellum force of B. subtilis was exponentially distributed.

Following the design stage of our project we moved on to the implementation stage. This involved construction of a cloning strategy, construction of our biobricks and transformation and characterisation of these biobricks in B. subtilis. For more information on this aspect of the project please see the Wet Lab Hub.

The testing and validation of our project can be split into three main areas;

• Work with B. subtilis - Including characterisation of growth curves and transformation,
• Characterisation and control of motility
• Production of Biomaterials in B. subtilis

Please see the Results Page for more information on the key results from the testing and validation.

Here is a summary of the achievements of the Imperial College 2008 team:

• Submitted 45 documented parts to the Registry
• Characterized and improved the existing part BBa J31005 (chloramphenicol acetyl transferase, CAT)
• Developed integration sequences for Biobricks, to allow devices to be constructed that can then be excised and planted into B. subtilis
• Layed the groundwork for future teams to work with B. subtilis by BioBricking promoters, RBSs, terminators and so on and characterising them
• Showed that expansion into other organisms is a definite possibility!
• Developed a method for tracking and analysing bacterial motility
• Helped Bristol by sending them a mini-iGEM project: Chemotactic dot-to-dot with information on quorum sensing and directed movement
• Helped Bristol by sending them part BBa_J37015 (AHL generator + GFP) from our 2007 stock which was an empty vector in the Registry

Of course, that is a very simplified description of our project. We expanded upon our project by looking into possible areas for real-world applications. For a case-study of such an implementation, check out how our project fits in with >>> Biocouture >>>