Team:KU Leuven/Project/DataPage

From 2013.igem.org

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   Welcome to our data page! Here we will summarize everything we achieved this summer. Of course, if you want a more extensive explanation you should check out the corresponding wiki page.
+
   Welcome to our data page! Here we will summarise everything we achieved this summer. Of course, if you want a more extensive explanation, please check out the corresponding wiki page.<br/>
 +
  Since the European Jamboree, we characterized our bricks further via <a href="https://2013.igem.org/Team:KU_Leuven/Project/Glucosemodel/MeS#Characterisation">Mass Spectrometry studies of MeS</a> and <a href="https://2013.igem.org/Team:KU_Leuven/Project/Glucosemodel/EBF#Characterisation">quantitated the aphid response to the EBF producing BanAphids</a>.  
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   <p align="justify">Ultimately our project aims to <b>reduce crop loss due to aphid infestations</b>. With an environmental project like ours, the computer is our best friend: through modelling and prediction algorithms we can reduce the real costs of field tests. Moreover, as iGEM team we were unable to conduct a field experiment for our BanAphids during this summer.<br/>
+
   <p align="justify">Our project aims to <b>reduce aphid infestations and thus improve crop yields for the industrial end-user and the private customer</b>. With an environmental project like ours, the importance of the computer and the feedback from our future end-users should not be underestimated. We adapted our project according to survey information and modelling results. Ultimately, we hope to reduce the costs of field tests via our <i>in silico</i> work.<br/><br/>
-
Our first step in the modelling was to <b>predict the effect of our pheromones on the environment and the ecosystem</b> through a series of modelling steps. You can find this in our <a href="https://2013.igem.org/Team:KU_Leuven/Project/Ecological/Modelling" target="_blank">ecological model page</a>.  
+
 
-
The pheromones E-β-farnesene and methyl salicylate will not only affect the environment but also the bacterial cell itself. To figure out the impact of our system on our <i>E. coli</i> we performed a <a href="https://2013.igem.org/Team:KU_Leuven/Project/Glucosemodel/MeS/Modelling-FBA" target="_blank">Flux Balance Analysis</a>.  Once we knew that our bacteria could handle the production of the pheromones we tried to <a href="https://2013.igem.org/Team:KU_Leuven/Project/Glucosemodel/MeS/Modelling" target="_blank">predict the exact production amounts</a> and find the rate limiting steps. Here our goal was also to feed wet-lab data into our algorithms. Finally, to optimise the impact of the released pheromones on the aphids and the ladybugs, we designed an <a href="https://2013.igem.org/Team:KU_Leuven/Project/Oscillator/Modelling" target="_blank">oscillating transcription factor network</a> to regulate their production. This oscillator also communicates between cells, enforcing the oscillating rhythm onto the whole colony.<br/>
+
First, we started our modelling on the <b><a href="https://2013.igem.org/Team:KU_Leuven/Project/Modelling/Cellular_Level" target="_blank">cellular level</a></b>. We must figure out the <b>impact of E-β-farnesene and methyl salicylate production on <i>E. coli</i></b>. Thus, we performed a <a href="https://2013.igem.org/Team:KU_Leuven/Project/Glucosemodel/MeS/Modelling-FBA" target="_blank">Flux Balance Analysis</a>. Results were compared with wetlab data such as growth curves and GC-MS data.
-
Summarized, <b>these algorithms allow us to model our system from the cellular metabolism throughout to the environmental impact</b>. Based on our models, we adapted the actual building of the system towards the most effective circuit. Finally, our mathematical predictions will provide significant benefits once we prepare our E. coligy for field tests.  
+
  We also aimed to <b><a href="https://2013.igem.org/Team:KU_Leuven/Project/Glucosemodel/MeS/Modelling" target="_blank">predict the exact amounts produced</a> and find the rate limiting steps</b>. Here we fed wetlab data into our algorithms. The outcome will define/defined our choice of promoters, plasmid copy number and additive requirements. </br></br>
-
  </p>
+
 
 +
Secondly we did a lot of modelling on the <b><a href="https://2013.igem.org/Team:KU_Leuven/Project/Modelling/Colony_Level">colony level</a></b>. We designed an <a href="https://2013.igem.org/Team:KU_Leuven/Project/Oscillator/Modelling" target="_blank">oscillating transcription factor network</a> to <b>regulate pheromone production for the "sticker enclosed" BanAphids</b>. This oscillator network allows communication between cells, enforcing a synchronised but oscillating production rhythm onto the whole colony. This will optimise the impact of our BanAphids on aphids and ladybugs even though a direct contact cue between aphids and BanAphids is prevented. We designed this model to answer the concerns of the private end-users regarding the spray (or honeydew) system.<br/><br/>
 +
 
 +
Finally, we must know the <b>effect of our pheromones on <a href="https://2013.igem.org/Team:KU_Leuven/Project/Modelling/Ecosystem_Level" target="_blank">the ecosystem</a></b>. We performed a series of modelling steps which you can find in our <a href="https://2013.igem.org/Team:KU_Leuven/Project/Ecological/Modelling" target="_blank">ecological model page</a>. This information is essential in several ways : </br>
 +
It will define the choice of pheromone production rate, which we can regulate through eg. promoter ranges.</br>
 +
Dispersion data will indicate the optimal spacing of the BanAphid stickers, key information for the end-user.<br/><br/>
 +
 
 +
Summarised, <b>these algorithms allow us to model our system from the cellular metabolism throughout to the environmental impact</b>. Based on our models, we continuously adapted the actual building of the system towards the most effective circuit. This will reduce costs and save time when our <b>BanAphids</b> are ready for field tests, and later for actual use.</p>
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  <h3 class="bg-green">Wetlab</h3></a>
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<p align="justify"> Our wetlab work consists of 3 experimental parts :</br>
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</p>
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<ol>
 +
<li> The production and testing of the <b><a href="https://2013.igem.org/Team:KU_Leuven/Project/Glucosemodel/MeS">methyl salicylate bricks</a></b></li>
 +
<li> The production and testing of the <b><a href="https://2013.igem.org/Team:KU_Leuven/Project/Glucosemodel/EBF">E-beta farnesene bricks</a></b></li>
 +
<li> <b><a href="https://2013.igem.org/Team:KU_Leuven/Project/Ecological/wetlab">Ecological work</a></b>, testing pheromone impact on the ecosystem (i.e. plants, ladybugs, ...). Here, we found <b>industrial partners</b> in the companies Biobest and pcfruit.</li>
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  <h3>Ecology</h3>
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  <h3>Methyl Salicylate Experiments</h3>
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   <p align="justify"><b>Two companies specialized in insects were very interested in our project. They invited us to perform experiments at their facilities.</b> These experiments proved that methyl salicylate not only attracts predators but also activates & protects the plant against aphids. They also proved that aphids are driven away by methyl salicylate.<br/>
+
   <p align="justify">
 +
Throughout the summer, we made <b>4 different BioBricks</b> involved in the production of methyl salicylate. After finding out that the MIT 2006 brick (<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_J45700" target="_blank">BBa_J45700</a>) only produced weak amounts of the wintergreen scent (MeS), we dove into the literature and discovered a possible lack of chorismate present in the bacteria. We tried to overcome this problem by overexpressing <i>aroG</i> in <i>E. coli</i>. In the literature, we found two mutations that could make DAHP synthase insensitive to allosteric inhibition. <b>We succeeded in biobricking the normal <i>aroG</i> gene</b>, which gives future teams the opportunity to introduce mutations themselves to overcome the chorismate problem. <b>We characterised our bricks with a renewed <a href="https://2013.igem.org/Team:KU_Leuven/Project/Glucosemodel/MeS#Smell Test">smell test</a>, <a href="https://2013.igem.org/Team:KU_Leuven/Project/Glucosemodel/MeS#Headspace GC">GC</a> and an <a href="https://2013.igem.org/Team:KU_Leuven/Project/Glucosemodel/MeS#SDS PAGE">SDS-PAGE</a> analysis.</b>
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  <h3>EBF project</h3>
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  <h3>(E)-β-farnesene Experiments</h3>
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   <p align="justify">After the whole summer's work, we finally made <b>5 Biobricks</b>. <br/>
+
   <p align="justify">After the whole summer's work, we finally made <b>5 BioBricks</b> for this section.
-
1. <a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1060001"BBa_K1060001</a> <br/>
+
Our favourites are:</p>
 +
<ol>
 +
<li><a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1060002" target="_blank">BBa_K1060002</a> contains an open reading frame that codes for (E)-β-farnesene synthase from <i>Artemisia annua</i>. The enzyme converts farnesyl diphosphate into E-β-farnesene. It was a milestone in our project work. <b>We succeeded to remove an <i>EcoRI</i> site in the gene (AY835398.1). This gave us one of the basic parts we needed to create our system</b>. </li>
 +
<li><a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1060009" target="_blank">BBa_K1060009</a> is a construct that constitutively expresses β-farnesene synthase. <b>This was the final device used for our</b> <a href="https://2013.igem.org/Team:KU_Leuven/Project/Glucosemodel/EBF" target="_blank">Aphid experiments</a>.</li>
 +
 +
 +
<li><a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1060011" target="_blank">BBa_K1060011</a> is similar to <a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1060009" target="_blank">BBa_K1060009</a>. However, in this biobrick we added a <i>lac</i> operator in front of the β-farnesene synthase. <b>This makes it possible to switch of (E)-β-farnesene production by using biosensors expressing LacI.</b></li>
 +
</ol>
 +
 +
<p align="justify">Our pilot studies with these biobricks and the <a href="https://2013.igem.org/Team:KU_Leuven/Project/Glucosemodel/EBF#aphid experiments">aphids</a> are promising and we now backed it up with quantitative data. Apart from this <i>in vivo</i> characterisation, we also initiated an <a href="https://2013.igem.org/Team:KU_Leuven/Project/Glucosemodel/EBF#SDSPAGE">SDS-PAGE</a> analysis.<b> Taken together this indicates that the EBF synthase is produced</b>.  </p>
-
The proof of concept experiments were successful. In the case of aphids experiments, the <a href=""video</a> showed the moving direction of aphids aligns with the the decreasing gradient of EBF concentration. On the other hand, the SDS-PAGE indicated that the EBF synthase was produced.
 
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<h3>Ecosystem Experiments</h3>
 +
  <p align="justify">Two companies (<b>Biobest and pcfruit</b>) specialised in biological pest management, were very <b>interested in our project</b> and invited us to perform experiments at their facilities. <b><a href="https://2013.igem.org/Team:KU_Leuven/Project/Ecological/wetlab" target="_blank">These experiments</a> demonstrated the effect of MeS in inducing plant defence mechanisms and that this has an effect on the aphid population</b>. <br/>
 +
</div>
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</div>
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   <a href="https://2013.igem.org/Team:KU_Leuven/Human_Practices">
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  <h3 class="bg-red">Human Practices</h3></a>
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  <p align="justify">As a result of our <b>unique collaboration between philosophers and scientists within our team, we formulated several new approaches to ethical and philosophical considerations</b>. The best example is the <a href="https://2013.igem.org/Team:KU_Leuven/Human_Practices/Ethics/Descriptive" target="_blank"> descriptive evaluation</a> in which all team members and supervisors were interviewed about their ethical beliefs. This bottom-up collaboration between students is a new starting point for approaching human practices within the iGEM competition. We also explained why participants should inform the general public using the ideas of philosopher <a href="https://2013.igem.org/Team:KU_Leuven/Human_Practices/HannahArendt" target="_blank">Hannah Arendt</a> and we examined the responsability of synthetic biologists using the ideas of philosopher <a href="https://2013.igem.org/Team:KU_Leuven/Human_Practices/HansJonas" target="blank">Hans Jonas</a>. <b>This is not only a first time in the iGEM competition, but has also never been done in the scientific literature!</b><br/>
+
  <p align="justify">As a result of our <b>unique collaboration between philosophers and scientists within our team</b>, we formulated a new additional approach: <b>Bottom-up or ethics from within</b>. This approach improves the communication between different stakeholders and achieves reciprocal understanding in early stages of the project. From this approach the government, scientists, the public and even the industry can benefit. This new approach is bottom-up structured where the dialogue between philosophers, scientists and the general public is central. We have developed a new and strong framework for this approach in which we also explained why we should inform the general public using the ideas of philosopher <a href="https://2013.igem.org/Team:KU_Leuven/Human_Practices/HannahArendt" target="_blank">Hannah Arendt</a> and we examined the responsibility of synthetic biologists using the ideas of philosopher <a href="https://2013.igem.org/Team:KU_Leuven/Human_Practices/HansJonas">Hans Jonas</a>. <b>This is not only a first time in the iGEM competition that these well-known philosophers are read to provide a strong foundation, but has also never been done in the scientific literature!</b><br/>
-
<p align="justify">To make sure that we have <a href="https://2013.igem.org/Team:KU_Leuven/Human_Practices/EndUser" target="blank">a product that is worth to be launched</a> we asked Biobest & pcfruit why they were so interested in the BanAphids. <b><br/><br/>Furthermore, we also asked farmers whether they would use our BanAphids and the results were definitely positive!</b></p>
+
In short, we consider this bottom-up collaboration between students a new starting point for approaching human practices within the iGEM competition.<br/>
 +
</p>
</p>
</p>
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</div>
 
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   <p align="justify"><b>We organized a symposium for the general public for which we also invited the other BeNeLux teams to present their project.</b> We hope that one day this will become a yearly tradition in the iGEM competition.<br/><br/>
+
   <p align="justify">
-
  We also went to <b>schools</b> to teach them about synthetic biology. For this we made our very own Plexiglas "biobricks" which the students can use to work on excercises and we made a <b>3D-bacterial model</b>, which gives the students an idea of what a bacterium looks like.</p>
+
Thanks to extensive dialogue we designed a product that interests both relevant industries like Biobest & pcfruit and the public. <b>Furthermore, we also asked <a href="https://2013.igem.org/Team:KU_Leuven/Human_Practices/EndUser">end users</a> (farmers) if they would use our BanAphids and the results were very positive!</b><br/><br/>
 +
 
 +
<b>Our symposium for the general public</b> was another opportunity for interaction with the public. <b>We invited the other Benelux (Belgium, The Netherlands, Luxembourg) teams to present their project.</b> We hope that this symposium will become a yearly tradition in the iGEM competition.<br/><br/>
 +
 
 +
We also go the youth interested in synthetic biology! We went to <b>high schools</b> with our very own <a href="https://static.igem.org/mediawiki/2013/8/84/Plexiglas_system.JPG"><b>Plexiglas "biobricks"</b></a> which the students can use to work get familiar with the BioBrick system and we made a <a href="https://static.igem.org/mediawiki/2013/5/57/3D_bacterial_model.JPG" target="_blank"><b>3D-bacterial model</b></a>, which gives the students an idea of what a bacterium looks like.</p>
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Latest revision as of 03:43, 29 October 2013

iGem

Secret garden

Congratulations! You've found our secret garden! Follow the instructions below and win a great prize at the World jamboree!


  • A video shows that two of our team members are having great fun at our favourite company. Do you know the name of the second member that appears in the video?
  • For one of our models we had to do very extensive computations. To prevent our own computers from overheating and to keep the temperature in our iGEM room at a normal level, we used a supercomputer. Which centre maintains this supercomputer? (Dutch abbreviation)
  • We organised a symposium with a debate, some seminars and 2 iGEM project presentations. An iGEM team came all the way from the Netherlands to present their project. What is the name of their city?

Now put all of these in this URL:https://2013.igem.org/Team:KU_Leuven/(firstname)(abbreviation)(city), (loose the brackets and put everything in lowercase) and follow the very last instruction to get your special jamboree prize!

tree ladybugcartoon

Welcome to our data page! Here we will summarise everything we achieved this summer. Of course, if you want a more extensive explanation, please check out the corresponding wiki page.
Since the European Jamboree, we characterized our bricks further via Mass Spectrometry studies of MeS and quantitated the aphid response to the EBF producing BanAphids.

Our project aims to reduce aphid infestations and thus improve crop yields for the industrial end-user and the private customer. With an environmental project like ours, the importance of the computer and the feedback from our future end-users should not be underestimated. We adapted our project according to survey information and modelling results. Ultimately, we hope to reduce the costs of field tests via our in silico work.

First, we started our modelling on the cellular level. We must figure out the impact of E-β-farnesene and methyl salicylate production on E. coli. Thus, we performed a Flux Balance Analysis. Results were compared with wetlab data such as growth curves and GC-MS data. We also aimed to predict the exact amounts produced and find the rate limiting steps. Here we fed wetlab data into our algorithms. The outcome will define/defined our choice of promoters, plasmid copy number and additive requirements.

Secondly we did a lot of modelling on the colony level. We designed an oscillating transcription factor network to regulate pheromone production for the "sticker enclosed" BanAphids. This oscillator network allows communication between cells, enforcing a synchronised but oscillating production rhythm onto the whole colony. This will optimise the impact of our BanAphids on aphids and ladybugs even though a direct contact cue between aphids and BanAphids is prevented. We designed this model to answer the concerns of the private end-users regarding the spray (or honeydew) system.

Finally, we must know the effect of our pheromones on the ecosystem. We performed a series of modelling steps which you can find in our ecological model page. This information is essential in several ways :
It will define the choice of pheromone production rate, which we can regulate through eg. promoter ranges.
Dispersion data will indicate the optimal spacing of the BanAphid stickers, key information for the end-user.

Summarised, these algorithms allow us to model our system from the cellular metabolism throughout to the environmental impact. Based on our models, we continuously adapted the actual building of the system towards the most effective circuit. This will reduce costs and save time when our BanAphids are ready for field tests, and later for actual use.

Our wetlab work consists of 3 experimental parts :

  1. The production and testing of the methyl salicylate bricks
  2. The production and testing of the E-beta farnesene bricks
  3. Ecological work, testing pheromone impact on the ecosystem (i.e. plants, ladybugs, ...). Here, we found industrial partners in the companies Biobest and pcfruit.

Methyl Salicylate Experiments

Throughout the summer, we made 4 different BioBricks involved in the production of methyl salicylate. After finding out that the MIT 2006 brick (BBa_J45700) only produced weak amounts of the wintergreen scent (MeS), we dove into the literature and discovered a possible lack of chorismate present in the bacteria. We tried to overcome this problem by overexpressing aroG in E. coli. In the literature, we found two mutations that could make DAHP synthase insensitive to allosteric inhibition. We succeeded in biobricking the normal aroG gene, which gives future teams the opportunity to introduce mutations themselves to overcome the chorismate problem. We characterised our bricks with a renewed smell test, GC and an SDS-PAGE analysis.

(E)-β-farnesene Experiments

After the whole summer's work, we finally made 5 BioBricks for this section. Our favourites are:

  1. BBa_K1060002 contains an open reading frame that codes for (E)-β-farnesene synthase from Artemisia annua. The enzyme converts farnesyl diphosphate into E-β-farnesene. It was a milestone in our project work. We succeeded to remove an EcoRI site in the gene (AY835398.1). This gave us one of the basic parts we needed to create our system.
  2. BBa_K1060009 is a construct that constitutively expresses β-farnesene synthase. This was the final device used for our Aphid experiments.
  3. BBa_K1060011 is similar to BBa_K1060009. However, in this biobrick we added a lac operator in front of the β-farnesene synthase. This makes it possible to switch of (E)-β-farnesene production by using biosensors expressing LacI.

Our pilot studies with these biobricks and the aphids are promising and we now backed it up with quantitative data. Apart from this in vivo characterisation, we also initiated an SDS-PAGE analysis. Taken together this indicates that the EBF synthase is produced.

Ecosystem Experiments

Two companies (Biobest and pcfruit) specialised in biological pest management, were very interested in our project and invited us to perform experiments at their facilities. These experiments demonstrated the effect of MeS in inducing plant defence mechanisms and that this has an effect on the aphid population.

As a result of our unique collaboration between philosophers and scientists within our team, we formulated a new additional approach: Bottom-up or ethics from within. This approach improves the communication between different stakeholders and achieves reciprocal understanding in early stages of the project. From this approach the government, scientists, the public and even the industry can benefit. This new approach is bottom-up structured where the dialogue between philosophers, scientists and the general public is central. We have developed a new and strong framework for this approach in which we also explained why we should inform the general public using the ideas of philosopher Hannah Arendt and we examined the responsibility of synthetic biologists using the ideas of philosopher Hans Jonas. This is not only a first time in the iGEM competition that these well-known philosophers are read to provide a strong foundation, but has also never been done in the scientific literature!
In short, we consider this bottom-up collaboration between students a new starting point for approaching human practices within the iGEM competition.

Thanks to extensive dialogue we designed a product that interests both relevant industries like Biobest & pcfruit and the public. Furthermore, we also asked end users (farmers) if they would use our BanAphids and the results were very positive!

Our symposium for the general public was another opportunity for interaction with the public. We invited the other Benelux (Belgium, The Netherlands, Luxembourg) teams to present their project. We hope that this symposium will become a yearly tradition in the iGEM competition.

We also go the youth interested in synthetic biology! We went to high schools with our very own Plexiglas "biobricks" which the students can use to work get familiar with the BioBrick system and we made a 3D-bacterial model, which gives the students an idea of what a bacterium looks like.