Nanopore analysis of nucleic acids
Butler, Thomas, 1978-
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Nanopore analysis of nucleic acids is a novel technique to study the physical properties of nucleic acids at the single-molecule level. It also has the potential to be a central component of a fundamentally new DNA sequencing methodology. In this technique, individual single-stranded DNA or RNA molecules are detected and characterized by an ionic current blockade produced when they are electrophoretically driven through a nanometer-scale pore. The primary focus of this thesis is a detailed characterization and interpretation of the sub-level structure of the ionic current blockades produced during the interaction between single-stranded DNA and RNA molecules and the Staphylococcus aureus alpha-Hemolysin protein nanopore. These investigations reveal important mechanistic details of the interaction between nucleic acids and alpha-Hemolysin. The results presented in this thesis contribute significantly to the growing utility of nanopore analysis as a model system to study biologically relevant physical and chemical processes, and they may help with the development of novel, nanopore-based technological applications. A secondary focus of this thesis is the initial steps toward genetically engineering Mycobacterium smegmatis porin A as an alternative protein nanopore for nucleic acid analysis. Successful creation of an additional protein nanopore represents a substantial scientific and technological breakthrough.
- Physics