Team:UC Chile2/Cyanolux/Project

From 2012.igem.org

(Difference between revisions)
Line 78: Line 78:
<br />
<br />
-
<p>We decided that we would separate LuxAB (the luciferase part of the operon) and LuxCDEG (the substrate producing enzymes LuxCDE with LuxG the FMNH2/FMN reducing enzyme) to allow phase-dependent expression of the parts. Using specific promoters of Synechocystis PCC. 6803 we can have fine-tunning of the production of bioluminescence. Recent work on global gene expression in Synechocystis aided on finding adecuate promoters[[#7 7]], [[#8 8]] . (Images at right from cited papers.)
+
<p>We decided that we would separate LuxAB (the luciferase part of the operon) and LuxCDEG (the substrate producing enzymes LuxCDE with LuxG the FMNH2/FMN reducing enzyme) to allow phase-dependent expression of the parts. Using specific promoters of Synechocystis PCC. 6803 we can have fine-tunning of the production of bioluminescence. Recent work on global gene expression in Synechocystis aided on finding adecuate promoters[[#7| 7]], [[#8| 8]] . (Images at right from cited papers.)
To try our approach, we selected various promoter which could serve the purpose.  Our rational for selecting candidate promoters involved amplitude of oscillation, peak activity, hour, absence of restriction sites, predicted strength of promoter according to the role of the gene and reproducibility between experiments (based on the literature available).  We looked for promoters which would have peak expression nearby dusk hours and that were slightly out of phase to optimize production of bioluminescence according to our mathematical models <b>(LINK OVER HERE!)</b>. We prioritized promoters from genes that would be involved in central energetic metabolism as we thought that their expression would be most robust and reliable. We choose the transaldolase promoter <b>(specific name here and code in Synechocystis Genome)</b> to direct the expression of the LuxAB genes and we found a couple of other promoters which filled the other requirements from above. Pcaa3 (NAME HERE AND DESCRIPTION OF ENDOGENOUS ACTIVITY) and PsigE (NAME HERE AND DESCRIPTION OF ENDOGENOUS ACTIVITY), the former being already in Biobrick format (courtesy from the Utah team iGEM 2010).
To try our approach, we selected various promoter which could serve the purpose.  Our rational for selecting candidate promoters involved amplitude of oscillation, peak activity, hour, absence of restriction sites, predicted strength of promoter according to the role of the gene and reproducibility between experiments (based on the literature available).  We looked for promoters which would have peak expression nearby dusk hours and that were slightly out of phase to optimize production of bioluminescence according to our mathematical models <b>(LINK OVER HERE!)</b>. We prioritized promoters from genes that would be involved in central energetic metabolism as we thought that their expression would be most robust and reliable. We choose the transaldolase promoter <b>(specific name here and code in Synechocystis Genome)</b> to direct the expression of the LuxAB genes and we found a couple of other promoters which filled the other requirements from above. Pcaa3 (NAME HERE AND DESCRIPTION OF ENDOGENOUS ACTIVITY) and PsigE (NAME HERE AND DESCRIPTION OF ENDOGENOUS ACTIVITY), the former being already in Biobrick format (courtesy from the Utah team iGEM 2010).

Revision as of 18:44, 22 September 2012

Project: Luxilla - Pontificia Universidad Católica de Chile, iGEM 2012