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Team:DTU-Denmark/test front
From 2014.igem.org
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$(this).find(".front-column-text").stop().css("height","0").animate({height: textHeight+"px"}, 600).parents(".front-page-curtain").css("box-shadow","0 0 10px black"); | $(this).find(".front-column-text").stop().css("height","0").animate({height: textHeight+"px"}, 600).parents(".front-page-curtain").css("box-shadow","0 0 10px black"); | ||
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Latest revision as of 17:52, 15 October 2014
- Home
- Overview
- Methods
- Achievements
- Team
At the Giant Jamboree, we received a gold medal and an Interlab Study Award. We also won the Overgrad Best Parts Collection.
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Problem:How can we quantify promoter activity?
When characterising promoters today it is done by proxy of protein, this can result in unwanted variations caused by translation and folding efficiency or excess cellular stress. Another important issue is the lack of a standard for measurements of fluorescence as most characterisations are done using fluorescent proteins such as GFP. As a result, most characterisations are done in relative units making it complicated or impossible to compare measurements between labs, strains and growth conditions. READ MORE (LINK to background) 
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Solution:Spinach RNA allows us to measure RNA concentration using fluorometry
The Spinach RNA is an aptamer, which can bind to DFHBI to create a fluorophore that resembles GFP in spectral properties. This provides a way to easily quantify RNA concentration and allows promoter activities to be calculated in units Polymerases Per Second. (Modelling link). The possibility of measuring promoter activity in absolute terms will enable researchers to share and compare results obtained in different labs, and better characterise the function of promoters. READ MORE (LINK to experimental design) 
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Results:We have developed a method that can be used to measure absolute promoter activity
Using RNA transcribed in vitro we created a standard series which can be used to determine the concentration of Spinach given a fluorescence intensity. We also designed an experiment to determine the degradation rate of the Spinach RNA in vivo. Combining these two experiments with the growth rate and fluorescence of a Spinach-expressing culture it is possible to determine the promoter activity using. READ MORE (LINK to results) 
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