Exploring RhoBAST Aptamer-Dye Complexes for Enhanced Metabolite Sensing

Undergraduate Honors Thesis

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Citeable URL: https://scholar.colorado.edu/concern/undergraduate_honors_theses/f4752j38q

Abstract

RNA is an essential component of cellular processes from gene expression to protein formation. A biosensor is a tool that turns binding of a metabolite of interest into a fluorescence signal. This project aims to explore and optimize RNA-based biosensors by comparing the binding affinity and fluorescence intensity between a previously researched RhoBAST-MaP555 aptamer-dye complex and other forms of rhodamine dyes. By evaluating the fluorescence output of JFX 549-Halotag and JF 549-alkyne, I aim to broaden the dyes that can be used in a RhoBAST complex. In future work this should increase the ability to track multiple metabolites with biosensors. The structural differences in deuterated JFX 549-Halotag and JF 549-alkyne, compared to MaP555 will lead to more insight on the structural necessities when it comes to the RhoBAST-dye complex. This thesis is a systematic examination of the steps necessary to acquire RhoBAST DNA, purified RNA, and Fluorescence Assays with reflections on my research experience as a whole.

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