Team:Evry/Sensor

From 2013.igem.org

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<h2>Construction of the sensor system</h2>
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<h2>Construction of the iron-responsive biosensors</h2>
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<i>E. coli</i>'s genome is composed of many Fur binding site. Based on a genome study, we identified 4 promoters which are controled by the FUR protein.
<i>E. coli</i>'s genome is composed of many Fur binding site. Based on a genome study, we identified 4 promoters which are controled by the FUR protein.
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<li>AceB promoter</li>
<li>AceB promoter</li>
<li>Fes promoter</li>
<li>Fes promoter</li>

Revision as of 17:33, 28 October 2013

Iron coli project

Iron Sensor


Construction of the iron-responsive biosensors

E. coli's genome is composed of many Fur binding site. Based on a genome study, we identified 4 promoters which are controled by the FUR protein.

  • AceB promoter
  • Fes promoter
  • FecA promoter
  • yncE promoter

We constructed iron-responsive biosensors by combining 3 genetic parts: an E. coli promoter with a Ferric Uptake Regulator (Fur) binding site, a fluorescent reporter (sfGFP), and a transcriptional terminator. These sensors respond to ambient iron by using the Fur system to repress a target gene.

We constructed 4 differents iron sensor using promoters regions from aceB (BBa_K1163102), fes (BBa_K1163108), fepA (BBa_K1163105) and yncE (BBa_K1163111). Finally, pAceB appears to be the best candidate to build our sensor system. See our results



Fig 1 Diagram of our genetic iron sensor. Iron binds the Ferric Uptake Regulator (Fur) to form a complex with high affinity for the Fur box in the promoter, here shown as the aceB promoter. Once the iron-Fur complex is bound to the promoter, it represses transcription of the target gene GFP. GFP expression is thus negatively correlated with iron availability.



Fig 2 Construction of an iron-responsive genetic element by fusing a Fur-regulated promoter with a reporter gene. Promoter-reporter fusions were made with flanking restriction sites that are compatible with Biobrick-based cloning.

NAME FIGURE DESCRIPTION

E. coli promoter with Fur binding site

iron-Fur complex binds promoter to repress expression

sfGFP

Fluorescent reporter gene

Terminator

terminator to stop transcription

Plasmid

Biobrick-compatible plasmid backbone

Table I Genetic elements used to make iron-responsive sensors.