Publication Date

2017-07-29

Availability

Embargoed

Embargo Period

2019-07-29

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PHD)

Department

Biochemistry and Molecular Biology (Medicine)

Date of Defense

2017-07-06

First Committee Member

Sapna K. Deo

Second Committee Member

Zafar Nawaz

Third Committee Member

Sanjoy K. Bhattacharya

Fourth Committee Member

Marcello Mascini

Abstract

The importance of nucleic acids detection to biological science has expanded as new technology enables historically unprecedented insight into their form and function. Additional challenges to assay development have resulted from the need to detect everything from enormous genomic sequences to miniscule non-coding RNA. The focus of this work centers on a recognition of the complexity required of modern analytical techniques and a strategy to eliminate as much technical surplus as can be achieved without detrimental performance. Initially, a laboratory-based technique has been developed to directly detect miRNA in complex matrices using fluorescence, and this was applied to quantitation of miR-155 in breast cancer samples. In order to provide a better dynamic range and limit of detection, a bioluminescence-based solid-phase platform was designed around a “bridged” capture system, for which detection of synthetic miR-34b was demonstrated. In addition, a novel fusion construct consisting of tamavidin 2 and Gaussia luciferase was created for use as a universal reporter. Finally, the requirement for laboratory infrastructure was abandoned completely in the development of a paper-based, point-of-care assay for the Zika virus. In summation, the work presented was integrated as part of future projects, both in progress and in planning, to provide readers with a framework for further development.

Keywords

nucleic acid detection; miRNA; point-of-care; bioluminescence

Available for download on Monday, July 29, 2019

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