Team:Toulouse/Project/Spreading

From 2014.igem.org

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<p class="texte">
<p class="texte">
It is known that endophyte bacteria must sporulate to survive to winter. In order to limit the spreading of our bacterium, we decided to limit its lifespan to only one season. The bacteria should be injected in spring, grow during the summer and finally should be inactivated in fall.<br\>  
It is known that endophyte bacteria must sporulate to survive to winter. In order to limit the spreading of our bacterium, we decided to limit its lifespan to only one season. The bacteria should be injected in spring, grow during the summer and finally should be inactivated in fall.<br\>  
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<i>Bacillus subtilis</i> is a sporing bacterium : sporulation enable the microorganism to handle very harsh conditions and to spread tree to tree. Indeed, a spore is a very resistant form that is adapted for unfavorable conditions and for dispersal.<br/>  
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<i>Bacillus subtilis</i> is a sporing bacterium: sporulation enable the microorganism to handle very harsh conditions and to spread tree to tree. Indeed, a spore is a very resistant form that is adapted for unfavorable conditions and for dispersal.<br/>  
To keep the control on the development of SubtiTree, our strain should therefore be non-sporing. We chose a <I>B.subtilis</I> strain without late genes of sporulation. Thus, after a season of treatment, the sap become less nutritious, the temperature is low and the engineered bacterium cannot survive the following winter.<br/>
To keep the control on the development of SubtiTree, our strain should therefore be non-sporing. We chose a <I>B.subtilis</I> strain without late genes of sporulation. Thus, after a season of treatment, the sap become less nutritious, the temperature is low and the engineered bacterium cannot survive the following winter.<br/>
In addition, deleting all the engineered bacterial community every year puts a brake on the evolution due to random mutation, thus it allows to keep control on the genetic constructions.
In addition, deleting all the engineered bacterial community every year puts a brake on the evolution due to random mutation, thus it allows to keep control on the genetic constructions.
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<p class="title1">Gene transfer: toxin-antitoxin system</p>
<p class="title1">Gene transfer: toxin-antitoxin system</p>
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<p class="texte"> While we were trying to respond to bacterial spreading problem, we also wondered about horizontal gene transfer.The goal of this module is to prevent horizontal transfer between bacteria. Indeed, it is necessary to avoid any exchange of genetic material between wild type organisms and optimized organisms : it could be dangerous because of mutations, and considering ethics, it seems to be essential to avoid the spreading of synthetic genes.
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<p class="texte"> While we were trying to respond to bacterial spreading problem, we also wondered about horizontal genes transfer. The goal of this module is to prevent horizontal transfers between bacteria. Indeed, it is necessary to avoid any exchange of genetic material between wild type organisms and optimized organisms: it could be dangerous because of mutations, and considering ethics, it seems to be essential to avoid the spreading of synthetic genes.
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<br>Considering this issue, we thought about a system to avoid such transfers : a toxin-antitoxin module. It involves the addition of two genes to the bacterium : a gene encoding for a toxin (for example <i>tse2</i>) and a gene encoding for the antitoxin (<i>tsi1</i>), placing them in an opposite way on the genome. The large space between them prevents simultaneous transfers : if the optimized bacterium transfers the gene encoding for the toxin, the probability that the gene encoding for the antitoxin may be transferred simultaneously is really low since they are located far away from each other.<br/>
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<br>Considering this issue, we thought about a system to avoid such transfers: a toxin-antitoxin module. It involves the addition of two genes to the bacterium: a gene encoding for a toxin (for example <i>tse2</i>) and a gene encoding for the antitoxin (<i>tsi1</i>), placing them in an opposite way on the genome. The large space between them prevents simultaneous transfers: if the optimized bacterium transfers the gene encoding for the toxin, the probability that the gene encoding for the antitoxin may be transferred simultaneously is really low since they are located far away from each other.<br/>
Therefore, if the host bacterium receives the gene encoding for the toxin, it will be unable to survive since it will not have the antitoxin. If it receives the antitoxin only, it will not be useful for the bacterium, and will not affect it.<br/>
Therefore, if the host bacterium receives the gene encoding for the toxin, it will be unable to survive since it will not have the antitoxin. If it receives the antitoxin only, it will not be useful for the bacterium, and will not affect it.<br/>
To sum up, since a simultaneous transfer is dimly probable, the bacterium will either die because of the toxin or live while expressing the antitoxin.  
To sum up, since a simultaneous transfer is dimly probable, the bacterium will either die because of the toxin or live while expressing the antitoxin.  
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<p class="title1">Using integrative plasmids</p>
<p class="title1">Using integrative plasmids</p>
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<p class="texte">
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All our constructions are carried by integrative plasmids. Consequently, our different genetic modules would be integrated into the bacterium genome. The integration in the genome is more stable as the constructions are less likely to be transferred to other microorganisms. In addition to that, the expression of our genetic modules would not be dependent on a selective pressure based on an antibiotic resistance (as we can not inject antibiotics in the tree), allowing a high level of transcription <i>in planta</i>.  
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All our constructions are carried by integrative plasmids. Consequently, our different genetic modules would be integrated into the bacterium genome. The integration in the genome is more stable as the constructions are less likely to be transferred to other microorganisms. In addition to that, the expression of our genetic modules would not be dependent on a selective pressure, allowing a high level of transcription <i>in planta</i>.  
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Revision as of 14:20, 14 October 2014