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==Light Sensing==
==Light Sensing==
[[Image:Imperial_2008_Holgram_Art.jpg | thumb | A holographic image of a work by sculptor Eileen Borgeson]]
[[Image:Imperial_2008_Holgram_Art.jpg | thumb | A holographic image of a work by sculptor Eileen Borgeson]]
The first step on the path to construction of a well-defined biomaterial shape [?] is guiding the microfactories into place and triggering production when they get there. To achieve this we need a stimulus - something that the bacteria can detect and respond to - that we can control accurately (possibly even in 3D!).
The first step on the path to construction of a well-defined biomaterial shape [?] is guiding the microfactories (an affectionate term for our bacteria) into place and triggering production when they get there. To achieve this we need a stimulus - something that the bacteria can detect and respond to - that we can control accurately (possibly even in 3D!).


The most obvious candidate for this is light. Light allows us to generate complex patterns with well defined edges, while gradients in wavelength and intensity will allow us to build up varying concentrations of biomaterial. After examining a number of light pathways present in ''subtilis'', other bacteria and light-sensing bricks in the Registry we decided to make use of a native pathway involving YtvA. This protein is used by ''B. subtilis'' to detect blue light, whereupon it triggers a cascade of interactions. Importantly some way down the chain a molecule called sigma B (σB) is produced which can act to boost the expression of genes.
The most obvious candidate for this is light. Light allows us to generate complex patterns with well defined edges, while gradients in wavelength and intensity will allow us to build up varying concentrations of biomaterial. After examining a number of light pathways present in ''subtilis'', other bacteria and light-sensing bricks in the Registry we decided to make use of a native pathway involving YtvA. This protein is used by ''B. subtilis'' to detect blue light, whereupon it triggers a cascade of interactions. Importantly some way down the chain a molecule called sigma B (σB) is produced which can act to boost the expression of genes.

Revision as of 03:23, 8 September 2008

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<html><a href=http://openwetware.org/wiki/IGEM:IMPERIAL/2008/Prototype><img width=50px src=http://openwetware.org/images/f/f2/Imperial_2008_Logo.png></img</a></html> Home The Project B.subtilis Chassis Wet Lab Dry Lab Notebook

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Authors: Wiki team

Editors: Wiki team

Chassis 1

This page offers a brief overview of how B. subtilis meets our main project specifications - to a much higher degree than E. coli! You'll find information on the main mechanisms behind our proposed system, and some very interesting biology too. If you would like even more depth than the outlines below, you are welcome to visit our OpenWetWare pages on those topics - but don't get to engrossed and forget to come back!

Light Sensing

A holographic image of a work by sculptor Eileen Borgeson

The first step on the path to construction of a well-defined biomaterial shape [?] is guiding the microfactories (an affectionate term for our bacteria) into place and triggering production when they get there. To achieve this we need a stimulus - something that the bacteria can detect and respond to - that we can control accurately (possibly even in 3D!).

The most obvious candidate for this is light. Light allows us to generate complex patterns with well defined edges, while gradients in wavelength and intensity will allow us to build up varying concentrations of biomaterial. After examining a number of light pathways present in subtilis, other bacteria and light-sensing bricks in the Registry we decided to make use of a native pathway involving YtvA. This protein is used by B. subtilis to detect blue light, whereupon it triggers a cascade of interactions. Importantly some way down the chain a molecule called sigma B (σB) is produced which can act to boost the expression of genes.

We plan to over-express YtvA, and use σB as a promoter for genes that stop movement and produce biomaterial. Thus when the bacteria detects blue light those genes will turn on, the bacteria will stop and biomaterial will be produced.

If we were really ambitious, or as a continuation of this project if it works well, one might try using holograms of blue light in semi-solid media to produce a 3D image that the microfactories could fill in with biomaterial!

Motility

A schematic of the B. subtilis rotary flagellar motor is shown. Motile cells are powered by interactions of the FliG protein with the MotA/B complex (which generates torque). The protein EpsE acts as a molecular clutch to disengage the rotary flagellar motor, leaving the flagellum intact but unpowered. This shuts down motility and facilitates biofilm formation. Fluorescence microscopy photos of B. subtilis show bacterial membranes in red and flagella in green. FliM and FliF are motor proteins [1]

To achieve accurate distribution of our biomaterial microfactories (an affectionate term for our bacteria) we need to be able to exert fine control over their motility. B. subtilis' prime method of getting about is flagellar locomotion; a ring of protein in the cell membrane rotates and is attached to a flagella that extrudes, acting like a long corkscrew propeller to push the cell through its environment.

Where B. subtilis differs from other bacteria is in our knowledge of its mechanism for this movement. A recent paper [LINK] described a clutch mechanism involved in the process; the flagella can be detached from the rotor by expression of a molecule that interacts with the flagella and distorts it so it is disengaged from the rotor protein.

Control over the expression of this protein should allow us very quick control of the bacteria - when we want it to stop we trigger expression of the clutch molecule which halts movement.

To draw a parallel with a car, current synthetic methods of stopping bacteria are akin to destroying the engine (which can take time!) in order to stop it powering the vehicle where the method we hope to take advantage of would be like putting the car into neutral - disengaging the engine from the driveshaft. It's an elegant solution that offers us quick control and also the opportunity for quick reversal (putting the car back into "drive").

More: http://openwetware.org/wiki/IGEM:IMPERIAL/2008/Prototype/Motility

Biomaterials

After our bacteria are positioned correctly, they need to be able to express a biomaterial. B. subtilis is Gram-positive, meaning it has only a single membrane as opposed to a double membrane like E. coli. This means that physical expression of a biomaterial is a lot more tractable; biomaterial can be produced and secreted more quickly, allowing a higher production efficiency. With a double membrane, material may collect inside the cell and destroy it from within.

Another important aspect of the biomaterial specifications is what we want to secrete; we did a lot of research on this and decided to try and produce elastin. Elastin is a small molecule (which means it can be produced and secreted more easily) that

http://openwetware.org/wiki/IGEM:IMPERIAL/2008/Prototype/Biomaterials



The next page details some more pros and cons about working with subtilis, as well as an overview of our development of it as a chassis... > The Second Chassis Page >


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