Team:LMU-Munich/Inverter

From 2012.igem.org

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<p align="justify">Last year, the LMU-Munich iGEM team designed a metal-sensing device. This device links metal sensing promoters to a visual output. However, it turned out that some of the promoters that respond to metal ions are negatively regulated. For this purpose, the LMU-Munich 2011 Team started a project to convert a positive input signal into a negative output or vice versa.</p>
<p align="justify">Last year, the LMU-Munich iGEM team designed a metal-sensing device. This device links metal sensing promoters to a visual output. However, it turned out that some of the promoters that respond to metal ions are negatively regulated. For this purpose, the LMU-Munich 2011 Team started a project to convert a positive input signal into a negative output or vice versa.</p>
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* To get an explanation of how it works and see the results: [[Team:LMU-Munich/Inverter#Theory and Results|Data and Results]]
* To get an explanation of how it works and see the results: [[Team:LMU-Munich/Inverter#Theory and Results|Data and Results]]
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* How to compose your individual Inverter, by fusions PCRs and 3a assemblies is explained here: [[Team:LMU-Munich/Inverter#Construct your own Inverter|Construct your own Inverter]]</p>
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* How to compose your individual Inverter, by fusions PCRs and 3A assemblies is explained here: [[Team:LMU-Munich/Inverter#Construct your own Inverter|Construct your own Inverter]]</p>
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<p align="justify">The main component of the [http://partsregistry.org/wiki/index.php?title=Part:BBa_K823040 Inverter] is the small RNA RyhB, which natively translationally inhibits the upstream fused region ''uof<sub>CGU</sub>'' of the ''fur'' gene by binding to it and therefore masking Shine Dalgarno sequence. </p>  
<p align="justify">The main component of the [http://partsregistry.org/wiki/index.php?title=Part:BBa_K823040 Inverter] is the small RNA RyhB, which natively translationally inhibits the upstream fused region ''uof<sub>CGU</sub>'' of the ''fur'' gene by binding to it and therefore masking Shine Dalgarno sequence. </p>  
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{| style="color:black;" cellpadding="3" width="70%" cellspacing="0" border="0" align="center" style="text-align:left;"
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<font color="#000000"; size="2"><p align="justify"> Fig. 1: Organization of fur mRNA and base-pairing between uof and RyhB. The uof and fur reading frames are depicted by white and black arrows, respectively. The sequence comprising nt −110 to +12 is enlarged. The putative SD sequences and start codons of uof and fur are boxed. The -GG- to -GGAGG- mutation in the putative SD of uof is indicated. </p></font>
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<font color="#000000"; size="2"><p align="justify"> '''Fig. 1: Organization of fur mRNA and base-pairing between uof and RyhB.''' The uof and fur reading frames are depicted by white and black arrows, respectively. The sequence comprising nt −110 to +12 is enlarged. The putative SD sequences and start codons of uof and fur are boxed. The -GG- to -GGAGG- mutation in the putative SD of uof is indicated. </p></font>
<!-- The uof codons UCA6 and AGA7 involved in iron-responsive decoding (see text) and the two consecutive stop codons of uof in the proximal-->
<!-- The uof codons UCA6 and AGA7 involved in iron-responsive decoding (see text) and the two consecutive stop codons of uof in the proximal-->
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<font color="#000000"; size="2"><p align="justify"> Fig. 2:Design and functional principle of the Inverter switch. </p></font>
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<font color="#000000"; size="2"><p align="justify"> '''Fig. 2: Design and functional principle of the Inverter switch.''' </p></font>
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<p align="justify">The β-galactosidase assay below shows the function of this Inverter. 1 mM IPTG is always present, so that the Reporter is fully induced. When grown with increasing amounts of arabinose in the medium, RyhB is produced and inhibits uof<sub>CGU</sub> and consequently the fused reporter in a concentration-dependent manner.</p>
<p align="justify">The β-galactosidase assay below shows the function of this Inverter. 1 mM IPTG is always present, so that the Reporter is fully induced. When grown with increasing amounts of arabinose in the medium, RyhB is produced and inhibits uof<sub>CGU</sub> and consequently the fused reporter in a concentration-dependent manner.</p>
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<p align="justify">In order to use the Inverter for the promoters and output of choice, it has to be constructed by fusion PCR (see next section).</p>
<p align="justify">In order to use the Inverter for the promoters and output of choice, it has to be constructed by fusion PCR (see next section).</p>
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{| style="color:black;" cellpadding="3" width="70%" cellspacing="0" border="0" align="center" style="text-align:left;"
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<font color="#000000"; size="2"><p align="justify"> Fig. 3: β-Galactosidase assay of Inverter with ''lacZα'' as reporter. The higher the arabinose concentration, the higher the translational repression of ''uof<sub>CGU</sub>-lacZα'' </p></font>
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<font color="#000000"; size="2"><p align="justify"> '''Fig. 3: β-Galactosidase assay of Inverter with ''lacZα'' as reporter.''' The higher the arabinose concentration, the higher the translational repression of ''uof<sub>CGU</sub>-lacZα'' </p></font>
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====Construct your own Inverter====
====Construct your own Inverter====

Latest revision as of 14:11, 26 October 2012

iGEM Ludwig-Maximilians-Universität München Beadzillus

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The LMU-Munich team is exuberantly happy about the great success at the World Championship Jamboree in Boston. Our project Beadzillus finished 4th and won the prize for the "Best Wiki" (with Slovenia) and "Best New Application Project".

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