Team:Valencia/Project

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Overview
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<h4><u>Synechosunshine</u>: photosynthetically powered biolamp </h4>
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        <ul>
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        <li><a href="https://2012.igem.org/Team:Valencia/design">Experimental Design</a></li>
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        <li><a href="https://2012.igem.org/Team:Valencia/results">Results and discussion</a></li>
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        <li><a href="https://2012.igem.org/Team:Valencia/Safety">Safety</a></li>
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        <li><a href="https://2012.igem.org/Team:Valencia/Acknowledgments">Acknowledgments</a></li>
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Our main goal for iGEM 2012 is the creation of a <strong>self-sufficient biolamp powered by solar light</strong>. In order to do this we have taken advantage of an artificial consortium between two naturally coexisting microorganisms: <strong><i>Synechococcus elongatus</i></strong> PCC 7942 and <strong><i>Aliivibrio fischeri</i></strong>. <i>A. fischeri</i> is a <strong>marine heterotrophic bacterium</strong>, capable of producing bioluminescence after activation by quorum sensing signals, while <i>S. elongatus</i> is a <strong>photosynthetic cyanobacterium</strong> that will allow the system to be fed by solar light.</div>
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We have arranged an <strong>artificial symbiotic interaction</strong> (communication and feeding) between these two organisms (cocultured in common broth separated by a semipermeable membrane) by the means of genetic engineering. The reason behind the biphasic setting is to avoid interference in the light emission-absorption between the different cultures to <strong>enhance the general energetic efficiency</strong> of the system. <br><br>
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<i>A. fischeri</i>’s genes for bioluminescence are regulated by an <strong>operator which is only activated at high concentrations of AHL</strong> (Acyl-homoserine-lactone), an own secreted quorum sensing molecule which is abundantly present at high population densities (aggregated in colonies, biofilms, host glands or phycospheres). When this bacterium lives free in open waters it does not express bioluminescence (Miyashiro and Ruby, 2012).<br><br>
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<center><a href="https://static.igem.org/mediawiki/2012/7/74/Definitivo_noche_y_d%C3%ADa.png"><img src="https://static.igem.org/mediawiki/2012/9/93/E4ea72b3edbf2ee66cccb588b372245b.gif" alt=""title="click to see it bigger!"></a></center>
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To <strong>induce bioluminescence</strong> in our system, we have modified <i>S. elongatus</i> to <strong>synthesize and export AHL</strong> to the common broth, in order to induce bioluminescence in <i>A. fischeri</i>. As our biolamp only requires to be ‘switched on’ at night, we are preceding the gene expressing AHLase enzyme (luxI) with a photosensitive operator. The genetic construct is based on the cyanobacterial <strong>promotor <i>psbAI</i></strong>, which responds to light through the molecular physiology of the cyanophyte photosystem (Golden, 1995). As we require an inverse response to light stress, we annexed the <a href="http://partsregistry.org/Part:BBa_Q04510" target="_blank">cI lambda inverter</a>, before ligating the luxI gene.<br><br>
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An important part of our project was to use the <strong>photosynthetic organism as a chemical energy donor</strong> for the luminescent heterotrophic population. To achieve this, we used a strain of <i>S. elongatus</i> developed by Ducat et al. (2012) from Harvard University which was transformed with a gene that expresses a transporter protein (cscB) to export sucrose to the culture medium in the presence of salt. This is how we render our biolamp self-sustainable, as the <strong>energy captured from the sun by <i>S. elongatus</i> is exported so that <i>A. fischeri</i> can use it</strong>.<br><br></p>
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<h4><u>Synechosunshine: photosynthetically powered biolamp</u></h4>
 
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Our main goal for iGEM 2012 is the creation of an autosufficient biolamp powered by solar light. In order to do this we have taken advantage of an artificial consortium between two naturally coexisting microorganisms: <i>Synechococcus elongatus</i> PCC 7942 and <i>Aliivibrio fischeri</i>. <i>A. fischeri</i> is a marine heterotrophic bacterium, capable of producing bioluminescence after activation by quorum sensing signals, while <i>S. elongatus</i> is a photosynthetic cyanobacterium that will allow the system to be fed by solar light.<br><br>
 
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We chose <i>Synechococcus</i> because of its great photosynthetic capacity due to its size and alometry, even though it is difficult to transform due to its complicated genome (it has 8 different plasmids with unrepeated genes!!). (foto nuestra coccus)<br><br>
 
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On the other hand, <i>Aliivibrio</i> is a gammaproteobacterium which has stablished symbiosis with other marine organisms, exchanging bioluminescence for glucidic nutrients. The best known example for this is the bobtail squid <i>Euprymna scolopes</i>, which uses <i>Aliivibrio</i> grown in ventral photophores for nocturnal counterillumination. Coevultion of <i>A. fischeri</i> with other species is very habitual.<br><br>
 
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We have arranged an artificial symbiotic interaction (communication and feeding) between these two organisms (cocultured in common broth separated by a semipermeable membrane) by the means of genetic engineering. The point of making a biphasic setting is to avoid interference in the light emission-absorption between the different cultures to enhance the general energetic efficiency of the system. (foto de aliivibrio brillando)<br><br>
 
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<i>A. fischeri</i>’s genes for bioluminescence are regulated by an operator which only activates at high concentrations of AHL (Acyl-homoserine-lactone), an autosecreted quorum sensing molecule which occurs abundantly at high population densities (aggregated in colonies, biofilms, host glands or phycospheres). When it lives free in the water it does not express bioluminescence.<br><br>
 
-
To induce bioluminescence in our system, we have modified <i>S. elongatus</i> to synthesize and export AHL to the common broth, in order to induce bioluminescence in <i>A. fischeri</i>. As our biolamp only requires to be ‘switched on’ at night, we are preceding the gene expressing AHLase enzyme (luxI) with a photosensitive operator. The genetic construct is based on the cyanobacterial promotor psbAI, which responds to light through the molecular physiology of the cyanophyte photosystem. As we require an inverse response to light stress, we annexed the <a href="http://partsregistry.org/Part:BBa_Q04510" target="_blank">cI lambda inverter</a>, before ligating the luxI gene.<br><br>
 
-
An important part of our project was to use the photosynthetic organism as a chemical energy donor for the luminescent heterotrophic population. To achieve this, we used a strain of <i>S. elongatus</i> developed by Ducat et al. (2010) from Harvard University, which was transformed with a gene which expresses a transporter protein (cscB) to export sucrose to the culture medium in the presence of salt. This is how we render our biolamp autosustainable, as the energy captured from the sun by <i>S. elongatus</i> is exported so that <i>A. fischeri</i> can use it.<br><br></p>
 
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<h4><u>Other Ideas</u></h4>
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Other parallel objective included transforming <i>Chlamydomonas reinhardtii</i> with a DNA gun to introduce the whole lux operon from Aliivibrio in the chloroplast’s genome, to render it bioluminescent.  The transformation of an eukaryotic autotroph could be the first step towards bioluminescent vascular plants, which could be easily used as environmentally friendly street lighting, despite being in theory less efficient than our symbiotic system, due to their robust physiognomy and little optimization for luminescent and photosynthetic functions. On the other hand, plants would be a more handy option, due to their environmental resistance and independence of a careful and constant upkeep. </p>
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<br><br><br>
<b>References</b>
<b>References</b>
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Ducat, D. C., Avelar-Rivas, A. J., Way, J. C. & Silver, P. A. (2012) Rerouting carbon flux to enhance photosynthetic productivity.<i> Applied and Environmental Microbiology</i>, 78(8):2660–2668.
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<br><br>
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Golden, S. S. (1995) Light-Responsive Gene Expression in Cyanobacteria. <i> J. Bacteriology</i>, 177(7): 1651-1654.
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<br><br>
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Miyashiro, T. & Ruby G. E. (2012) Shedding light on bioluminiscence regulation in Vibrio fischeri. <i>Mol. Micro</i>., 84(5):795-806.
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Latest revision as of 03:51, 27 September 2012



Overview


Synechosunshine: photosynthetically powered biolamp


http://www.flickr.com. Edquint8364

Our main goal for iGEM 2012 is the creation of a self-sufficient biolamp powered by solar light. In order to do this we have taken advantage of an artificial consortium between two naturally coexisting microorganisms: Synechococcus elongatus PCC 7942 and Aliivibrio fischeri. A. fischeri is a marine heterotrophic bacterium, capable of producing bioluminescence after activation by quorum sensing signals, while S. elongatus is a photosynthetic cyanobacterium that will allow the system to be fed by solar light.

We have arranged an artificial symbiotic interaction (communication and feeding) between these two organisms (cocultured in common broth separated by a semipermeable membrane) by the means of genetic engineering. The reason behind the biphasic setting is to avoid interference in the light emission-absorption between the different cultures to enhance the general energetic efficiency of the system.

A. fischeri’s genes for bioluminescence are regulated by an operator which is only activated at high concentrations of AHL (Acyl-homoserine-lactone), an own secreted quorum sensing molecule which is abundantly present at high population densities (aggregated in colonies, biofilms, host glands or phycospheres). When this bacterium lives free in open waters it does not express bioluminescence (Miyashiro and Ruby, 2012).

To induce bioluminescence in our system, we have modified S. elongatus to synthesize and export AHL to the common broth, in order to induce bioluminescence in A. fischeri. As our biolamp only requires to be ‘switched on’ at night, we are preceding the gene expressing AHLase enzyme (luxI) with a photosensitive operator. The genetic construct is based on the cyanobacterial promotor psbAI, which responds to light through the molecular physiology of the cyanophyte photosystem (Golden, 1995). As we require an inverse response to light stress, we annexed the cI lambda inverter, before ligating the luxI gene.

An important part of our project was to use the photosynthetic organism as a chemical energy donor for the luminescent heterotrophic population. To achieve this, we used a strain of S. elongatus developed by Ducat et al. (2012) from Harvard University which was transformed with a gene that expresses a transporter protein (cscB) to export sucrose to the culture medium in the presence of salt. This is how we render our biolamp self-sustainable, as the energy captured from the sun by S. elongatus is exported so that A. fischeri can use it.






References

Ducat, D. C., Avelar-Rivas, A. J., Way, J. C. & Silver, P. A. (2012) Rerouting carbon flux to enhance photosynthetic productivity. Applied and Environmental Microbiology, 78(8):2660–2668.

Golden, S. S. (1995) Light-Responsive Gene Expression in Cyanobacteria. J. Bacteriology, 177(7): 1651-1654.

Miyashiro, T. & Ruby G. E. (2012) Shedding light on bioluminiscence regulation in Vibrio fischeri. Mol. Micro., 84(5):795-806.

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