Difference between revisions of "Part:BBa K1758376"
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This part contains all features from [https://parts.igem.org/Part:BBa_K1758102 BBa_K1758102] plus the blc-operator site described in [http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4083662/ Pan et al. 2013]. BlcR (see [https://parts.igem.org/Part:BBa_K1758370 K1758370]) can bind to the operator site.   
 
This part contains all features from [https://parts.igem.org/Part:BBa_K1758102 BBa_K1758102] plus the blc-operator site described in [http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4083662/ Pan et al. 2013]. BlcR (see [https://parts.igem.org/Part:BBa_K1758370 K1758370]) can bind to the operator site.   
 
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For more detailed information, please be referred to <a href="https://parts.igem.org/Part:BBa_K1758377">K1758377</a>
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For more detailed information, please be referred to <a href="https://parts.igem.org/Part:BBa_K1758377">K1758377</a>.
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<span class='h3bb'>Sequence and Features</span>
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<partinfo>BBa_K1758376 SequenceAndFeatures</partinfo>
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<h2> Usage and Biology</h2>
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<p>The reporter part was used to build a biosensor detecting ingredients of date rape drugs, namely GBL and GHB that also functions in cell free protein synthesis (CFPS) assay. Upon induction of T7 polymerase, sfGFP is produced. The following operator site from <i>Agrobacterium tumefaciens</i> is the binding site for the repressor BlcR. Our designed, translation enhancing 5'-untranslated region (5'-UTR; <a href="https://parts.igem.org/Part:BBa_K1758100">BBa_K1758100</a>) enhances sfGFP production. In presence of a the repressor BlcR(<a href="https://parts.igem.org/Part:BBa_K1758370">BBa_K1758370</a>) the reporter gene translation is weakened. BlcR weakens expression as long as no GHB or GBL is supplemented.</p>
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<h2> Results </h2>
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<p> Sequences of this part was used in <i>in vivo</i> as well as in <i>in vitro</i> experiments. This part in particular was used as a control for our <i>in vivo</i> experiments regarding our GHB / GBL biosensor device, as depicted in the following figure.</p>
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<figure>
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<a href="https://static.igem.org/mediawiki/2015/0/01/Bielefeld-CeBiTec_Blc_in_vivo_characterization.png"><img src="https://static.igem.org/mediawiki/2015/0/01/Bielefeld-CeBiTec_Blc_in_vivo_characterization.png" style="width:700px"></a>
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<figcaption> <i>In vivo characterization of GBL / GHB sensor with strain containing BBa_K1758377. All experiments were perfomed as triplicates. All samples except "control, not induced" were induced to express T7 polymerase at OD600 = 0.7-0.8 </i>. Control refers to a strain carrying this part in pSB1C3. </figcaption>
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</figure>
  
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<h2>Biology</h2>
 
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<p>The reporter part was used to build a biosensor detecting ingredients of date rape drugs, namely GBL and GHB that also functions in cell free protein synthesis (CFPS) assay. The promoter part (T7) is inducible by rhamnose addition. The following operator site from <i>Agrobacterium tumefaciens</i> is the binding site for the repressor BlcR. The specific 5' untranslated region enhances translation. sfGFP was used as reporter protein. In presence of a the repressor BlcR(<a href="https://parts.igem.org/Part:BBa_K1758370">BBa_K1758370</a>) the reporter gene translation is inhibited. In presence of additional GHB/GHB the repression is resolved and the reporter gene is translated.</p>
 
 
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<span class='h3bb'>Sequence and Features</span>
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<partinfo>BBa_K1758376 SequenceAndFeatures</partinfo>
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Revision as of 19:33, 19 September 2015

GHB / GBL biosensor - reporter part

This part contains all features from BBa_K1758102 plus the blc-operator site described in [http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4083662/ Pan et al. 2013]. BlcR (see K1758370) can bind to the operator site. For more detailed information, please be referred to K1758377. Sequence and Features BBa_K1758376 SequenceAndFeatures

Usage and Biology

The reporter part was used to build a biosensor detecting ingredients of date rape drugs, namely GBL and GHB that also functions in cell free protein synthesis (CFPS) assay. Upon induction of T7 polymerase, sfGFP is produced. The following operator site from Agrobacterium tumefaciens is the binding site for the repressor BlcR. Our designed, translation enhancing 5'-untranslated region (5'-UTR; BBa_K1758100) enhances sfGFP production. In presence of a the repressor BlcR(BBa_K1758370) the reporter gene translation is weakened. BlcR weakens expression as long as no GHB or GBL is supplemented.

Results

Sequences of this part was used in in vivo as well as in in vitro experiments. This part in particular was used as a control for our in vivo experiments regarding our GHB / GBL biosensor device, as depicted in the following figure.

In vivo characterization of GBL / GHB sensor with strain containing BBa_K1758377. All experiments were perfomed as triplicates. All samples except "control, not induced" were induced to express T7 polymerase at OD600 = 0.7-0.8 . Control refers to a strain carrying this part in pSB1C3.