Overview
During 2014 iGEM competition, teams were requested to analyse the efficiency of 3 different genetic devices (BioBricks) using GFP as a marker of gene expression. Here, iGEM ITESM CEM team presents the results of this interlab fluorescence measurement study.
The three devices analysed are composed by a variable promoter, a gene encoding a mutant Green Fluorescence Protein (GFP) used as a marker of expression, and a plasmid backbone. Two promoters (BBa_J23101 and BBa_J23115, recently renamed BBa_K823005 and BBa_K823012 at iGEM’s catalogue) are used, both of them being members of a family of constitutive promoters described by Chris Anderson, member of iGEM Berkley Team, in 2006 (1). This family of parts is registered at the catalogue under the alphanumeric codes BBa_J23100 – BBa_J23119.
Two different plasmid backbones are used: a low-copy (psB3K3) and a high-copy plasmid (psB1C3). The GFP-expressing BioBrick remains the same for all devices (registered at the catalog as BBa_E0240), and is composed of a ribosome binding site (RBS), a mutant GFP gene, and two termination sequences.
The aim of this study is to report the relative efficiency of the following genetic devices:
1) Promoter BBa_K823005 in low-copy plasmid psB3K3
2) Promoter BBa_K823005 in high-copy plasmig psB1C3
3) Promoter BBa_K823012 in high-copy plasmid psB1C3
In order to do so, GFP (BBa_E0240) is used as a marker of gene expression or reporter gene, because of the ease of fluorescence measurement experiments.
GFP has long been used as a reporter of patterns of gene expression in both prokaryotes, were it is useful for characterization of promoters, enhancers and terminators; and in eukaryotes, were tissue-specific or time-specific gene expression can be traced (2). The basis of this procedure is the usage of GFP’s fluorescence as a reporter of activity of promoters and enhancers; the relative fluorescence of cells at different experimental conditions can be compared with statistical techniques, and so the efficiency of the parts can be tested.
Existing GFP device
Bla bla.
New GFP device 1
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New GFP device 2
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