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Title: Synthetic biology biosensor design for medical diagnostics
Author: Kylilis, Nicolas
ISNI:       0000 0004 8504 4028
Awarding Body: Imperial College London
Current Institution: Imperial College London
Date of Award: 2017
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Biosensors are compact, integrated analytical devices that use a biologically derived sensor element to specifically detect their target analyte. Their design makes them convenient for the performance of diagnostic tests at the site where the sample is procured. Synthetic biology has been extensively applied to biosensor design to produce genetically encoded devices that detect metal and organic contaminants, drug molecules and metabolites among others. However, there is only a very limited number of synthetic biology biosensor tools available for the detection of analytes that do not readily transport across the cell membrane. This is especially the case for protein analytes despite the potential of such tools for use in diagnostic applications at the point of care. This research investigated synthetic biology biosensor designs for the detection of protein analytes and developed such technologies featuring high specificity sensor elements. One of the technologies developed features a single domain antibody (nanobody) which is surface displayed on E. coli cells. By the use of a GFP binding nanobody, this biosensor technology was successfully demonstrated to detect dimeric GFP in a plate agglutination assay format. A second technology developed features a RNA-aptamer sensor element incorporated into a RNA-switch device. For prototyping this technology, a thrombin binding RNA-aptamer incorporated into the RNA-switch device was demonstrated to detect thrombin by controlling GFP expression in E. coli cell-extract assay. Additionally, the design space for both prototyped technologies was successfully investigated for strategies that would enable the modulation of their analytical characteristics. Lastly, the modularity of the biosensor designs was investigated by the incorporation of sensors for alternative protein analytes. The pool of known biomarkers for medical conditions includes a large number of protein molecules. Since the prototyped designs feature sensor elements that can be evolved in vitro for the specific recognition of any protein target at will, the technologies developed here can potentially be utilised as synthetic biology platforms for the development of low cost biosensor devices for use in medical diagnostic applications.
Supervisor: Freemont, Paul ; Polizzi, Karen Sponsor: Biotechnology and Biological Sciences Research Council ; NetScientific (Firm)
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral