Difference between revisions of "Team:Bielefeld-CeBiTec"

Line 26: Line 26:
 
<!--<div class="row">-->
 
<!--<div class="row">-->
 
     <center>
 
     <center>
−
<img class="picture" src="https://static.igem.org/mediawiki/2016/0/03/Bielefeld_CeBiTec_2016_10_13_X_Evobodies_animation.gif" >
+
<img style="heigth:100px;" class="picture" src="https://static.igem.org/mediawiki/2016/0/03/Bielefeld_CeBiTec_2016_10_13_X_Evobodies_animation.gif" >
 
</center>
 
</center>
 
<br>
 
<br>
Line 71: Line 71:
 
</a>
 
</a>
 
</div>-->
 
</div>-->
 +
<br>
 
<div class="row">
 
<div class="row">
 
<div class="col-md-12 ">
 
<div class="col-md-12 ">
Line 86: Line 87:
 
<div class="thumbnail">  
 
<div class="thumbnail">  
 
<div class="row equal" id="top_left">
 
<div class="row equal" id="top_left">
−
<div class="col-xs-7 col-md-7 ">  
+
<div class="col-xs-3 col-md-3 ">  
 
<figure class="figure">
 
<figure class="figure">
 
<a href="https://2016.igem.org/Team:Bielefeld-CeBiTec/Project/Library"><img  class="picture figure-image" src="https://static.igem.org/mediawiki/2016/c/c9/Bielefeld_CeBiTec_2016_10_18_LIB_project_overview_library.png" /> </a>
 
<a href="https://2016.igem.org/Team:Bielefeld-CeBiTec/Project/Library"><img  class="picture figure-image" src="https://static.igem.org/mediawiki/2016/c/c9/Bielefeld_CeBiTec_2016_10_18_LIB_project_overview_library.png" /> </a>
 
</figure>
 
</figure>
 
</div>
 
</div>
−
<div class="col-xs-5 col-md-5">
+
<div class="col-xs-9 col-md-9">
 
<h3 class="textHeadline"><a href="https://2016.igem.org/Team:Bielefeld-CeBiTec/Project/Library"
 
<h3 class="textHeadline"><a href="https://2016.igem.org/Team:Bielefeld-CeBiTec/Project/Library"
 
>Library</a></h3>
 
>Library</a></h3>
Line 103: Line 104:
 
<div class="thumbnail">  
 
<div class="thumbnail">  
 
<div class="row equal" id="top_right">
 
<div class="row equal" id="top_right">
−
<div class="col-xs-7 col-md-7">  
+
<div class="col-xs-3 col-md-3">  
 
<a href="https://2016.igem.org/Team:Bielefeld-CeBiTec/Project/Mutation"><img  class="picture" src="https://static.igem.org/mediawiki/2016/d/d4/Bielefeld_CeBiTec_2016_10_14_project_description_mutation.png" />
 
<a href="https://2016.igem.org/Team:Bielefeld-CeBiTec/Project/Mutation"><img  class="picture" src="https://static.igem.org/mediawiki/2016/d/d4/Bielefeld_CeBiTec_2016_10_14_project_description_mutation.png" />
 
</div>
 
</div>
−
<div class="col-xs-5 col-md-5">
+
<div class="col-xs-9 col-md-9">
 
<h3 class="textHeadline"> <a href="https://2016.igem.org/Team:Bielefeld-CeBiTec/Project/Mutation">Mutation</a> </h3>
 
<h3 class="textHeadline"> <a href="https://2016.igem.org/Team:Bielefeld-CeBiTec/Project/Mutation">Mutation</a> </h3>
 
<p class="stdText"><a href="https://2016.igem.org/Team:Bielefeld-CeBiTec/Mutation">Overview</a><br><a href="https://2016.igem.org/Team:Bielefeld-CeBiTec/Results/Mutation">Results</a></p>
 
<p class="stdText"><a href="https://2016.igem.org/Team:Bielefeld-CeBiTec/Mutation">Overview</a><br><a href="https://2016.igem.org/Team:Bielefeld-CeBiTec/Results/Mutation">Results</a></p>
Line 118: Line 119:
 
<div class="thumbnail">  
 
<div class="thumbnail">  
 
<div class="row" id="botton_left">
 
<div class="row" id="botton_left">
−
<div class="col-xs-7 col-md-7">  
+
<div class="col-xs-3 col-md-3">  
 
<a href="https://2016.igem.org/Team:Bielefeld-CeBiTec/Project/Selection/Motivation"><img class="picture" src="https://static.igem.org/mediawiki/2016/9/9e/Bielefeld_CeBiTec_2016_10_14_project_description_selection.png" /></a>
 
<a href="https://2016.igem.org/Team:Bielefeld-CeBiTec/Project/Selection/Motivation"><img class="picture" src="https://static.igem.org/mediawiki/2016/9/9e/Bielefeld_CeBiTec_2016_10_14_project_description_selection.png" /></a>
 
</div>
 
</div>
Line 154: Line 155:
 
<td style="border:none; width:20px"><img src="https://static.igem.org/mediawiki/2016/4/48/Bielefeld_CeBiTec_2016_10_18_XXXX_tick.png" class="check" width="40px"></td>
 
<td style="border:none; width:20px"><img src="https://static.igem.org/mediawiki/2016/4/48/Bielefeld_CeBiTec_2016_10_18_XXXX_tick.png" class="check" width="40px"></td>
 
<td style="text-align:left; border:none;">
 
<td style="text-align:left; border:none;">
−
Design and construction of a <a href="https://2016.igem.org/Team:Bielefeld-CeBiTec/Project/Library/Overview" target="_blank">Evobody library</a> and invention of a new BioBrick class  </td>
+
Design and construction of an <a href="https://2016.igem.org/Team:Bielefeld-CeBiTec/Project/Library/Overview" target="_blank">Evobody library</a> and invention of a new BioBrick class  </td>
 
</tr>
 
</tr>
 
<tr style="border:none;">
 
<tr style="border:none;">
 
<td style="border:none; width:20px"><img src="https://static.igem.org/mediawiki/2016/4/48/Bielefeld_CeBiTec_2016_10_18_XXXX_tick.png" class="check" width="40px"></td>
 
<td style="border:none; width:20px"><img src="https://static.igem.org/mediawiki/2016/4/48/Bielefeld_CeBiTec_2016_10_18_XXXX_tick.png" class="check" width="40px"></td>
 
<td style="text-align:left; border:none;">
 
<td style="text-align:left; border:none;">
−
DeOver 100,000 clones per <a href="https://2016.igem.org/Team:Bielefeld-CeBiTec/Project/Library/Overview" target="_blank">library</a> generated  </td>
+
Over 100,000 clones per <a href="https://2016.igem.org/Team:Bielefeld-CeBiTec/Project/Library/Overview" target="_blank">library</a> generated  </td>
 
</tr>
 
</tr>
  
Line 166: Line 167:
 
<td style="border:none; width:20px"><img src="https://static.igem.org/mediawiki/2016/4/48/Bielefeld_CeBiTec_2016_10_18_XXXX_tick.png" class="check" width="40px"></td>
 
<td style="border:none; width:20px"><img src="https://static.igem.org/mediawiki/2016/4/48/Bielefeld_CeBiTec_2016_10_18_XXXX_tick.png" class="check" width="40px"></td>
 
<td style="text-align:left; border:none;">
 
<td style="text-align:left; border:none;">
−
High diversity confirmed by <a href="https://2016.igem.org/Team:Bielefeld-CeBiTec/Results/Library/Sequencing" target="_blank">high-throughput sequencing </a> </td>
+
High diversity of the plasmid library confirmed by <a href="https://2016.igem.org/Team:Bielefeld-CeBiTec/Results/Library/Sequencing" target="_blank">high-throughput sequencing </a> </td>
 
</tr>
 
</tr>
  
Line 179: Line 180:
 
<tr style="border:none;">
 
<tr style="border:none;">
 
<td style="border:none; width:20px"><img src="https://static.igem.org/mediawiki/2016/4/48/Bielefeld_CeBiTec_2016_10_18_XXXX_tick.png" class="check" width="40px"></td>
 
<td style="border:none; width:20px"><img src="https://static.igem.org/mediawiki/2016/4/48/Bielefeld_CeBiTec_2016_10_18_XXXX_tick.png" class="check" width="40px"></td>
−
<td style="text-align:left; border:none;">Construction, detailed characterization via high-throughput sequencing and submission of a plasmid-specific <a href="https://2016.igem.org/Team:Bielefeld-CeBiTec/Results/Mutation" target="_blank"> mutagenesis systems </a>  </td>
+
<td style="text-align:left; border:none;">Construction and detailed characterization via high-throughput sequencing of a plasmid-specific <a href="https://2016.igem.org/Team:Bielefeld-CeBiTec/Results/Mutation" target="_blank"> mutagenesis system </a>  </td>
 
</tr>
 
</tr>
  
Line 194: Line 195:
 
<td style="border:none; width:20px"><img src="https://static.igem.org/mediawiki/2016/4/48/Bielefeld_CeBiTec_2016_10_18_XXXX_tick.png" class="check" width="40px"></td>
 
<td style="border:none; width:20px"><img src="https://static.igem.org/mediawiki/2016/4/48/Bielefeld_CeBiTec_2016_10_18_XXXX_tick.png" class="check" width="40px"></td>
 
<td style="text-align:left; border:none;">
 
<td style="text-align:left; border:none;">
−
Plasmid sequence improvement by <a href="https://2016.igem.org/Team:Bielefeld-CeBiTec/Results/Mutation/Sequencing" target="_blank"> re-sequencing </a> and <a href="https://2016.igem.org/Team:Bielefeld-CeBiTec/Results/Mutation/Assembly" target="_blank"> <i>de novo</i> assembly reversion assays</a>   </td>
+
Plasmid sequence improvement by <a href="https://2016.igem.org/Team:Bielefeld-CeBiTec/Results/Mutation/Sequencing" target="_blank"> re-sequencing </a> and <i>de novo</i> assembly  </td>
 
</tr>
 
</tr>
  

Revision as of 03:11, 20 October 2016




We developed a novel and easy to use system for the generation of binding proteins in E. coli via in vivo directed evolution. Resulting proteins called Evobodies have the potential to bind specific to target proteins enabling varies medical and analytical applications. Big advantages of our low-cost system are the short hands on time and the short generation time.

As the starting point of our system, we designed and synthesized a library of a new entity of BioBricks encoding binding proteins in E. coli based on Nanobodies as well as Monobodies. The diversity of the library in E. coli is continuously increased by co-expressing a special DNA-Polymerase conferring plasmid restricted error-prone replication of the binding protein expressing plasmids. Finally, binding proteins with high affinity to the target protein are selected using a bacterial two-hybrid system providing growth advantage to antibiotics in relation to protein-protein interaction strength. Ultimately, desired clones are enriched during fermentation in batch or in continuous culture.

Binding capacities of our libraries, efficiency of our selection system and potential of our mutagenesis system were demonstrated. Moreover, library diversity and mutation system characteristics were analyzed in detail by high-throughput sequencing.



Achievements

Design and construction of an Evobody library and invention of a new BioBrick class
Over 100,000 clones per library generated
High diversity of the plasmid library confirmed by high-throughput sequencing
Functionality of the library was demonstrated by binding to various targets
Construction and detailed characterization via high-throughput sequencing of a plasmid-specific mutagenesis system
Construction and characterization of several parts for reversion assays
Plasmid sequence improvement by re-sequencing and de novo assembly
Construction and characterization of a bacterial two-hybrid selection system
Prediction of the outcome of our system
Evolution-based human practice projects were perfectly integrated
Concept development for industrial upscaling