Team:Uppsala University/Translational

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<td class="subtext"><h2>sRNA candidates</h2></td>
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<td class="subtext"><h2>Mechanism of downregulation</h2></td>
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<img src="https://static.igem.org/mediawiki/2012/7/74/Mechanism_of_downregulation.png" height="600" alt="Mechanism_of_downregulation">
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<b>The general system for downregulation is illustrated above.</b>
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<b>Stabilizing scaffold</b><br>
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In order to retain transcribed sRNA in E.coli and avoid degradation of the sRNA, a stabilizing scaffold can be linked to the domain which is hypothesized to interact with the domain of interest.
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<b>Hfq Protein</b><br>
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The Hfq Protein is a natively expressed protein which interacts with the scaffold. A deletion study of the Hfq protein resulted in total loss of regulation for many sRNAs [2].

Revision as of 16:18, 23 September 2012

Team Uppsala University – iGEM 2012


Silencing Truncated AAC(6') by use of fluorescent reporter

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In order to find candidates for conditional gene silencing, we used a method already described in [1] and adapted it to a modular biobrick RFC10 system. Briefly, the method goes like this:

  • Express a chimeric gene - The gene interest linked to a reporter in a strain
  • Express a library of small RNA's with a suitable stabilizing scaffold in the same strain
  • Screen for regulation of the reporter

Mechanism of downregulation

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Mechanism_of_downregulation


The general system for downregulation is illustrated above.
Stabilizing scaffold
In order to retain transcribed sRNA in E.coli and avoid degradation of the sRNA, a stabilizing scaffold can be linked to the domain which is hypothesized to interact with the domain of interest. Hfq Protein
The Hfq Protein is a natively expressed protein which interacts with the scaffold. A deletion study of the Hfq protein resulted in total loss of regulation for many sRNAs [2].

References

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[1] Sharma, V., Yamamura, A., Yokobayashi, Y., 2012. Engineering Artificial Small RNAs for Conditional Gene Silencing in Escherichia coli. ACS Synth. Biol. 1, 6–13.


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