NSTI Nanotech 2009

Highly specific anti-prothrombin aptamers

C-W Chi, Y-H Lao, L-C Chen
Department of Bio-Industrial Mechatronics Engineering, National Taiwan University, TW

Keywords: aptamer, prothrombin, SELEX, microarray

Abstract:

In vitro selection of anti-prothrombin aptamers is of great interest for anticoagulant drug development as well as early-stage thrombosis diagnosis. In this work, prothrombin was immobilized on magnetic beads for aptamer selection. After 10 rounds of selection, 60 clones were picked and 26 candidate sequences were identified. We chose 5 conserved sequences to test their specificity by an aptamer microarray assay. These aptamers have high specificity to prothrombin and do not crosstalk with thrombin. Analyzing the ratio of fluorescence intensity of prothromin to thrombin, we find that the signal of aptamer E2 is even fifty times higher than that of the reported anti-prothrombin aptamer F16. We also compared the GC contents of the initial ssDNA pool, 26 candidates, 5 conserved sequences, aptamer B8 and E2, and the reported aptamer F16. The result is implied that anti-prothrombin aptamers tends to have high GC content (up to 65.5%). Dissociation constants and functional assays of the anti-thrombin aptamers are under investigation and will be presented in the symposium along with the above findings.
 
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