IGEM:Paris Bettencourt 2012/Notebooks/Delay group

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=Alginate beads=
=Alginate beads=
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Here, we would place alginate beads, with controlled alginate matrix size into the capsule. The alginate beads contains theophylline, that would act as an activator of a [http://en.wikipedia.org/wiki/Riboswitch| riboswitch] that would repress gene expression [1]. This riboswitch would be fused into the 5'-UTR of the gene we want to repress (see figure 1). That gene would be one of the [http://en.wikipedia.org/wiki/Colicin| colicin] gene, that can diffuse and kill other ''Enterobacteriaceae''. The colicine gene is under Rpos promoter which is activated when bacteria is on stationary phase. So, bacteria  will die on stationary phase and when theophylline is under a certain threshold, thanks to its diffusion through beads and capsule.
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Here, we would place alginate beads, with controlled alginate matrix size into the capsule. The alginate beads contains theophylline, that would act as an activator of a [http://en.wikipedia.org/wiki/Riboswitch| riboswitch] that would repress gene expression [1]. On of our options is to deliver theophylline through bits of complex sugars that would be the carbon source of our bacteria. The riboswitch would be fused into the 5' end of the gene we want to repress (see figure 1). That gene would be one of the [http://en.wikipedia.org/wiki/Colicin| colicin] gene, that can diffuse and kill other ''Enterobacteriaceae''. The riboswitch will be translated, and thus we need to ensure that it will not affect the function of the protein. The colicine gene is under Rpos promoter which is activated when bacteria is on stationary phase (e.g. YiaGp). So, bacteria  will die on stationary phase and when theophylline is under a certain threshold, thanks to its diffusion through beads and capsule.
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However, we need to ensure that the expression of this toxin protein is very repressed and that it will not affect the fitness of the bacteria as long as it is in stationary phase with theophylline.
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If possible, we would like to try to replace the nuclease domain of the colicin gene with a restriction enzyme domain to target exclusively the synthetic genes carried by the plasmid.
[[Image:Theophuline_riboswitch.png|center|500px]]
[[Image:Theophuline_riboswitch.png|center|500px]]
<center>Figure 1 : Theophylline riboswitch, adapted from [1]</center>
<center>Figure 1 : Theophylline riboswitch, adapted from [1]</center>

Revision as of 11:11, 29 June 2012

Notebook Design Roadmap Meetings and to-dos Protocols Bibliography Previous Biosafety iGEM projects


Delay group

The aim of that subproject is to find a way to launch the destruction of either the construction or the organism after a certain laps of time.

Here we report the idea we have, and how we could make them real.

Alginate beads

Here, we would place alginate beads, with controlled alginate matrix size into the capsule. The alginate beads contains theophylline, that would act as an activator of a riboswitch that would repress gene expression [1]. On of our options is to deliver theophylline through bits of complex sugars that would be the carbon source of our bacteria. The riboswitch would be fused into the 5' end of the gene we want to repress (see figure 1). That gene would be one of the colicin gene, that can diffuse and kill other Enterobacteriaceae. The riboswitch will be translated, and thus we need to ensure that it will not affect the function of the protein. The colicine gene is under Rpos promoter which is activated when bacteria is on stationary phase (e.g. YiaGp). So, bacteria will die on stationary phase and when theophylline is under a certain threshold, thanks to its diffusion through beads and capsule. However, we need to ensure that the expression of this toxin protein is very repressed and that it will not affect the fitness of the bacteria as long as it is in stationary phase with theophylline. If possible, we would like to try to replace the nuclease domain of the colicin gene with a restriction enzyme domain to target exclusively the synthetic genes carried by the plasmid.

Figure 1 : Theophylline riboswitch, adapted from [1]

References

1. Topp, S. & Gallivan, J.P. Riboswitches in unexpected places -€” A synthetic riboswitch in a protein coding region. RNA journal 2498-2503 (2008) Paper

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