| Project/Area Number |
18560747
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Biofunction/Bioprocess
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| Research Institution | Tokyo University of Agriculture and Technology |
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Principal Investigator |
IKEBUKURO Kazunori Tokyo University of Agriculture and Technology, Institute of Symbiotic Science and Technology, Assoc. Prof. (70251494)
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| Project Period (FY) |
2006 – 2007
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| Project Status |
Completed (Fiscal Year 2007)
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| Budget Amount *help |
¥3,950,000 (Direct Cost: ¥3,500,000、Indirect Cost: ¥450,000)
Fiscal Year 2007: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000)
Fiscal Year 2006: ¥2,000,000 (Direct Cost: ¥2,000,000)
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| Keywords | aptamer / marker protein for disease / simultaneous screening / diagnosis technology / genetic algorithm / molecular recognition device / aptamer blotting / protein detection technology |
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| Research Abstract |
We have developed the screening method of aptamer based on target specificity using protein competitors. It is difficult to screen aptamers according to specificity by conventional SELEX, since such methods screen aptamers only based on their affinity. In our selection method, the library is simultaneously incubated with the target protein and other proteins used as competitors. We evaluated the screened library with the aptamer blotting method, which visualizes the binding of oligonucleotides to proteins. In this study, we identified two DNA aptamers against Tau protein and luciferase after only several rounds of selection by this method. The isolated aptamers had high affinity for the target protein, and did not bind to the competitor protein. These results suggest that our selection method may be useful for efficiently selecting specific aptamers against target proteins.
Moreover, we applied this method to screen the aptamers that bind to a specific uncharacterized protein in target tissue without purification. The conventional in vitro selection methods against proteins in tissue cannot be applied to the selection against uncharacterized proteins or it is not certain whether aptamers to target proteins are obtained during selection. We developed the selection method of aptamers to an uncharacterized protein in target tissue by visualizing the bindings of oligonucleotide to each protein in target tissue and isolating bound oligonucleotides from detected bands. This method would enable us to select aptamers to any protein in target tissue. We therefore believe that this method would be useful in selecting aptamers for analyses of uncharacterized proteins.
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