Team:Cambridge/Bioluminescence/Photinus pyralis

From 2010.igem.org

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We are investigating a number of firefly luciferases. As a comparison for our future experiments we will begin by cloning the existing Biobrick for [http://partsregistry.org/Part:BBa_I712019 wild type <i>P. pyralis</i> luciferase] into our strains of interest.
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The luciferase of the North American firefly, ''P. pyralis'', is a tried and tested mechanism for creating bioluminescence.  We were aware that a luciferase from this organism was already present in the registry ([http://partsregistry.org/Part:BBa_I712019 BBa_I712019]). We wanted to improve on this by three techniques:
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* [https://2010.igem.org/Team:Cambridge/Codons Codon optimisation] for expression in E. coli to increase the rate of translation
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* Using a mutant with increased substrate affinity
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* Parallel use of the Photinus pyralis [https://2010.igem.org/Team:Cambridge/Bioluminescence/Luciferin_Regeneration luciferin regenerating enzyme] to both relieve inhibition by oxyluciferin and increase availability of luciferin.
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Our project will focus on characterising both this existing luciferase and those we hope to submit to the registry:
 
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* An enhanced <i>P. pyralis</i> luciferase with increased brightness
 
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* A number of differently coloured versions of <i>L. cruciata</i> luciferase
 
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We intend to synthesise these luciferases on operons also containing the LRE for the firefly species in question, and then to quantify a PoPS-to-photons function.
 
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Revision as of 09:35, 18 October 2010