Short FRET: Accurate short distance measurements for synthetic biology and drug discovery.

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Democratising Single-molecule FRET and new fluorescence assays with sub 3 nm sensitivityThe key aims of this project are:1) To develop an open-source, economic, robust, and user-friendly instrument for single-molecule measurements, that anyone can build and use2) To develop new quantitative fluorescence assays sensitive to small changes in the structure of biomolecules (e.g. DNA, RNA and proteins) The student will be designing, building and validating a new instrument (a confocal microscope) capable of detecting individual fluorescent molecules, one at a time. Unlike commercial instruments in this class, this new instrument will not be based on a microscope body, and the student will work out how to provide all the necessary information for other groups to build the instrument. We anticipate this new instrument will cost a fraction of the price of commercial versions, and will, therefore, be an attractive alternative for many groups. A second objective will be to adapt a current fluorescence assay based on quenching (quenchable FRET) to develop this into a fully quantitative assay. This will involve molecular dynamics modelling, photo-physical characterisation, and ensemble and single-molecule experiments, to determine the mechanism of quenching and establish a relationship between quenching and biomolecular structure that can be used by us and others to observe small structural changes in biomolecules, which are inaccessible to current methods.