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Team:ZJU-China/Project/TheGhostKit/GhostSensor
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=== A universal platform for protein on-site detection === | === A universal platform for protein on-site detection === | ||
Interest in developing portable detection devices for on-site analysis has risen sharply in recent years. Many areas, including medical diagnostics, environmental toxicology and bioterrorism controls, would benefit from devices that can perform a rapid first-level detection without the need for equipped laboratories or skilled personnel. ZJU-China iGEM team this year successfully built a universal protein detection platform based on the idea of aptamer-induced inner-membrane scaffold dimerization. | Interest in developing portable detection devices for on-site analysis has risen sharply in recent years. Many areas, including medical diagnostics, environmental toxicology and bioterrorism controls, would benefit from devices that can perform a rapid first-level detection without the need for equipped laboratories or skilled personnel. ZJU-China iGEM team this year successfully built a universal protein detection platform based on the idea of aptamer-induced inner-membrane scaffold dimerization. | ||
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| + | In our project, we altogether used two sets of inner-membrane scaffolds as protein sensors. One is based on split GFP and the other is based on split β-lactamase. | ||
=== Split TEM-1 β-lactamase === | === Split TEM-1 β-lactamase === | ||
Revision as of 01:39, 29 October 2013
Contents |
Background
A universal platform for protein on-site detection
Interest in developing portable detection devices for on-site analysis has risen sharply in recent years. Many areas, including medical diagnostics, environmental toxicology and bioterrorism controls, would benefit from devices that can perform a rapid first-level detection without the need for equipped laboratories or skilled personnel. ZJU-China iGEM team this year successfully built a universal protein detection platform based on the idea of aptamer-induced inner-membrane scaffold dimerization.
In our project, we altogether used two sets of inner-membrane scaffolds as protein sensors. One is based on split GFP and the other is based on split β-lactamase.
Split TEM-1 β-lactamase
Protein fragment complementation assay is a widely used method for protein interaction detection. The two fragments of the protein is rational designed and based on protein interaction assisted folding. TEM-1 β-lactamase is relatively small and monomeric, is well characterized structurally and functionally, can be easily expressed, and is not toxic to prokaryotic cells. Furthermore, no orthologs of β-lactamase exist in many prokaryotes, and thus a protein fragment complementation assay based on β-lactamase could be used universally in prokaryotes without any intrinsic background activity. We split TEM-1 β-lactamase according to [4] into N-terminal consisting of a.a. 26-196 and C-terminal consisting of a.a. 198-290.
Diagram illustrating the split of TEM-1 β-lactamase.
(André Galarneau et al., Nature Biotechnology, 2002, 20, 619-22)
The activity of TEM-1 β-lactamase can be detected with a wide range of methods, such as simple colorimetric in vitro assays using the cephalosporin nitrocefin and assays in intact cells using the fluorescent substrate. There are also many commercial immune colloidal gold strips in China to detect β-lactamase in food and beverage. These strips are cheap, sensitive and robust, making them the ideal candidate for large scale applications.
