Date of Award

6-2008

Document Type

Union College Only

Degree Name

Bachelor of Science

Department

Biochemistry

First Advisor

Brian Cohen

Language

English

Keywords

fret, beacons, dye, fluorescent, oligonucleotides

Abstract

Quantum dots (QD) are fluorescent semiconductor nanostructures that have the ability to emit light through the visible and infrared regions. Due to various coatings of quantum dots, they can be conjugated to macromolecules such as peptides, DNA or RNA. Oligonucleotides tethered to fluorophores or quenchers can be used for such conjugations to achieve fluorescence resonance energy transfer (FRET) between the QD and the dye/quencher when excited by UV light; this is due to the hairpin sequences encoded into the nucleotides, which bring the FRET acceptor within the Foster radius of the QD. In order to utilize these molecular beacons as gene expression monitors, reversibility of FRET is necessary. This was achieved by incubation of beacons with oligonucleotides of complimentary sequence and allowing the strands to denature fully and anneal. An increase in the distance between the fluorescent dye and the quantum dot in beacons hybridized to complimentary oligonucleotides brought the dye out of the Foster radius and resulted in FRET reversal. FRET reversal was observed using steady state fluorescence analysis and fluorescent lifetime measurement. As a result of this study, a successful molecular beacon as well as quenched beacon conjugation protocol was established and reversible FRET was achieved with organic dyes as FRET acceptors through complementation.

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