Difference between revisions of "Team:TU Darmstadt/Composite Part"

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    <h1>COMPOSITE PART</h1>
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  <p><b>A genetic circuit is generated ensuring that genetically modified E.coli cannot survive outside of the required conditions. This genetic circuit is based on the availability of a non-natural amino acid (nnAA) which continuously has to be added to the medium. A reporter protein called mVenus is expressed as the level of nnAA in the medium decreases, signalizing the low nnAA concentration. If the level decreases further, colicin will no longer be inhibited, which leads to the complete destruction of the bacterias DNA and eventually their death. In order to keep the metabolic burden of our system as low as possible, we engineered a minimalized Colicin which consists of the DNase domain only. In addition to that, we employed a method to easily measure the metabolic burden by integrating GFP into the genome and even generated the possibility to integrate our entire safety approach into the genome to reduce the metabolic burden again.  
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One essential part in designing new BioBricks is the characterization and thus the estimation of the resources which are consumed by that BioBrick. For that reason, we wanted to genomically integrate a GFP coding gene into the genome of <i>E.coli</i> For the measurement of the metabolic burden caused by a BioBrick one can transform the cells with that BioBrick. After inducing the questioned BioBrick one can estimate the metabolic Burden by the decrease in constitutively expressed GFP.
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To make this kind of measurement possible we assembled this BioBrick, which is composed of the &lambda;&#8209;<i>attp</i>&#8209;Site, a J23101&nbsp;Promoter from the Anderson Promoter Series, a RBS and a coded GFP with an added LVA&#8209;Tag. The two key parts of this Brick are the &lambda;&#8209;<i>attp</i>&#8209;Site, which is necessary for the genomic integration of the Plasmid. The integration follows the &lambda;&#8209;bacteriophage integration mechanism combining the attachment site plasmid (<i>attP</i>) with the attachment site bacteria (<i>attB</i>). The second part is the constitutively expressed GFP with an LVA&#8209;Tag which makes the measurement of the metabolic burden possible and easily achievable.
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For making an easy and reliable method for a characterization of the metabolic burden caused by different BioBricks possible we think, that this part is worthy of achieving the price for the Best&nbsp;Composite&nbsp;Part.
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Revision as of 13:12, 19 October 2016

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iGEM TU Darmstadt 2016
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COMPOSITE PART

One essential part in designing new BioBricks is the characterization and thus the estimation of the resources which are consumed by that BioBrick. For that reason, we wanted to genomically integrate a GFP coding gene into the genome of E.coli For the measurement of the metabolic burden caused by a BioBrick one can transform the cells with that BioBrick. After inducing the questioned BioBrick one can estimate the metabolic Burden by the decrease in constitutively expressed GFP. To make this kind of measurement possible we assembled this BioBrick, which is composed of the λ‑attp‑Site, a J23101 Promoter from the Anderson Promoter Series, a RBS and a coded GFP with an added LVA‑Tag. The two key parts of this Brick are the λ‑attp‑Site, which is necessary for the genomic integration of the Plasmid. The integration follows the λ‑bacteriophage integration mechanism combining the attachment site plasmid (attP) with the attachment site bacteria (attB). The second part is the constitutively expressed GFP with an LVA‑Tag which makes the measurement of the metabolic burden possible and easily achievable. For making an easy and reliable method for a characterization of the metabolic burden caused by different BioBricks possible we think, that this part is worthy of achieving the price for the Best Composite Part.