Team:Imperial College London/Project/Switch/Overview

From 2011.igem.org

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<h1>Containment Device</h1>
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<h1>Preventing Horizontal Gene Transfer</h1>
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As part of our human practices work, we need to consider what will happen in the event that these bacteria are released into the soil. The potential consequences of their release relate to their uncontrolled spread and the possibility that they pass on the auxin genes to a potentially pathogenic bacterium.
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As part of our human practices work, we need to consider what will happen in the event that these bacteria are released into the soil. The potential consequences of their release relate to their uncontrolled spread and the possibility that they pass on the auxin genes to naturally occuring soil bacteria.
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The auxin that we are using is the natural indole-3-acetic acid, which is not used as a herbicide like many other synthetic auxins. However, in high concentrations, indole-3-acetic acid can retard growth - as shown in our experiments.  
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The auxin compound that we are using is the natural indole-3-acetic acid, which is not used as a herbicide like many other synthetic auxins. However, in high concentrations, indole-3-acetic acid can retard plant growth - as shown in our experiments.  
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While there are already a few species of bacteria that are able to secrete auxin<sup>[1]</sup>, it would be careless of us to release our bacteria without giving some thought to a containment device.
While there are already a few species of bacteria that are able to secrete auxin<sup>[1]</sup>, it would be careless of us to release our bacteria without giving some thought to a containment device.
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<h2>Kill Switch Designs</h2>
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<h2>The BacTrap Design</h2>
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The first idea was to implant a chemical into the selected region of soil that the bacteria would be unable to live without, but this was dismissed on environmental reasons as there is very little that we can add to the soil without damaging its composition or the organisms within.
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We have implemented the Holin/Anti-Holin regulated kill switch designed by the Berkeley 2008 iGEM team to create a system limiting horizontal gene transfer. Holin is a protein that forms pores in cell membranes and anti-holin binds to holin, inhibiting it's action. Once pores are formed by holin, lysozyme can access the periplasmic space and degrade the cell wall, causing cell lysis.  
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The second idea was to implement the Holin/Anti-Holin regulated kill switch designed by the Berkeley 2008 iGEM team under the control of a UV sensitive promoter. Since UV light can only penetrate 0.3mm into soil, this would be an ideal method to ensure that the bacteria remain underground.
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The third idea uses the same BioBrick parts from the 2008 Berkeley iGEM team to create a toxin/anti-toxin system to try and limit horizontal gene transfer. The Antiholin gene will be on the genome of the bacteria under the control of a strong promoter. The Holin and Lysozyme genes will be present on the same plasmid as the two auxin genes. The idea here is that the presence of the antiholin will prevent the cell from lysing from the effects of holin and lysozyme. In a different cell, i.e., one that does not have antiholin on its genome, the antiholin and lysozyme will kill the cell, preventing it from keeping the plasmid containing the auxin genes.
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In the BacTrap system, the antiholin gene will be on the genome of our engineered bacteria under the control of a strong promoter. The Holin and Lysozyme genes will be present on the same plasmid as the two auxin genes and the chemoreceptor gene. The idea here is that the presence of the antiholin will prevent the cell from lysing from the effects of holin and lysozyme. In a different cell, i.e., one that does not have antiholin on its genome, the antiholin and lysozyme will kill the cell, preventing it from keeping the plasmid containing the auxin genes. This mechanism will prevent the succesfull horizontal gene transfer to naturally occuring soil bacteria. <p>
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Revision as of 09:45, 18 August 2011




Preventing Horizontal Gene Transfer

As part of our human practices work, we need to consider what will happen in the event that these bacteria are released into the soil. The potential consequences of their release relate to their uncontrolled spread and the possibility that they pass on the auxin genes to naturally occuring soil bacteria.

The auxin compound that we are using is the natural indole-3-acetic acid, which is not used as a herbicide like many other synthetic auxins. However, in high concentrations, indole-3-acetic acid can retard plant growth - as shown in our experiments.

While there are already a few species of bacteria that are able to secrete auxin[1], it would be careless of us to release our bacteria without giving some thought to a containment device.


The BacTrap Design

We have implemented the Holin/Anti-Holin regulated kill switch designed by the Berkeley 2008 iGEM team to create a system limiting horizontal gene transfer. Holin is a protein that forms pores in cell membranes and anti-holin binds to holin, inhibiting it's action. Once pores are formed by holin, lysozyme can access the periplasmic space and degrade the cell wall, causing cell lysis.

In the BacTrap system, the antiholin gene will be on the genome of our engineered bacteria under the control of a strong promoter. The Holin and Lysozyme genes will be present on the same plasmid as the two auxin genes and the chemoreceptor gene. The idea here is that the presence of the antiholin will prevent the cell from lysing from the effects of holin and lysozyme. In a different cell, i.e., one that does not have antiholin on its genome, the antiholin and lysozyme will kill the cell, preventing it from keeping the plasmid containing the auxin genes. This mechanism will prevent the succesfull horizontal gene transfer to naturally occuring soil bacteria.


References

[1] http://m.biotecharticles.com/Biology-Article/Natural-Growth-Hormone-IAA-Indole-3-Acetic-Acid-602.html

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