The exploration of a quantum dot fluorescence resonance energy transfer (QD-FRET) based bioassay for label-free target nucleic acid detection is reported herein. This work explores the potential for developing a displacement assay for detection of nucleic acid sequences of various lengths, including one of 484 bases. Short probe oligonucleotides conjugated to QDs were allowed to hybridize to short partially mismatched dye-labelled oligonucleotide targets. The non-labelled target of interest, a 484-base segment of heat shock protein 70 (HSP 70), contained a portion that was fully complementary to the probe. Thermodynamic parameters suggested that HSP 70 would displace dye-labelled targets; however, detection was not observed. Modifications were made to this assay to reduce sterics and increase the stability of hybrids. The results obtained using this modified assay indicated that detection of non-labelled, long oligonucleotide sequences was possible using a displacement assay that relied on a short probe oligonucleotide.
| Identifer | oai:union.ndltd.org:TORONTO/oai:tspace.library.utoronto.ca:1807/33260 |
| Date | 20 November 2012 |
| Creators | Kamaluddin, Sara |
| Contributors | Krull, Ulrich Jorg |
| Source Sets | University of Toronto |
| Language | en_ca |
| Detected Language | English |
| Type | Thesis |
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