Abstract
BackgroundA Nanopore Detector provides a means to transduce single molecule events into observable channel current changes. Nanopore-based detection can report directly, or indirectly, on single molecule kinetics. The nanopore-based detector can directly measure molecular characteristics in terms of the blockade properties of individual molecules – this is possible due to the kinetic information that is embedded in the blockade measurements, where the adsorption-desorption history of the molecule to the surrounding channel, and the configurational changes in the molecule itself, imprint on the ionic flow through the channel. This rich source of information offers prospects for DNA sequencing and single nucleotide polymorphism (SNP) analysis. A nanopore-based detector can also measure molecular characteristics indirectly, by using a reporter molecule that binds to certain molecules, with subsequent distinctive blockade by the bound-molecule complex.ResultsIt is hypothesized that reaction histories of individual molecules can be observed on model DNA/DNA, DNA/Protein, and Protein/Protein systems. Preliminary results are all consistent with this hypothesis. Nanopore detection capabilities are also described for highly discriminatory biosensing, binding strength characterization, and rapid immunological screening.ConclusionIn essence, the heart of chemistry is now accessible to a new, single-molecule, observation method that can track both external molecular binding states, and internal conformation states.
Highlights
A Nanopore Detector provides a means to transduce single molecule events into observable channel current changes
1990's three direct measures of the force have been performed at the single molecule level: (1) the force required to break AT or GC base pairs [1,2,3], (2) the force required to extend single or double stranded DNA through distinct structural conformations, e.g., B form to S form DNA [4,5], etc., and (3) the forces exerted by polymerases working on polynucleotides [6]
A nanopore-based detector can directly measure molecular characteristics in terms of the blockade properties of individual molecules – this is possible due to the kinetic information that is embedded in the blockade measurements, where the adsorption-desorption history of the molecule to the surrounding channel, and the configurational changes in the molecule itself, imprint on the ionic flow through the channel [7,8,9,10,11,12]. (Note: the hypothesis that the current blockade patterns are caused by adsorption-desorption, and conformational flexing, is not conclusively proven, preliminary work on mechanism [11] and the success of the experimental approaches [7,8,9,10,11,12] add growing credence to this hypothesis.) This offers prospects for DNA sequencing and single nucleotide polymorphism (SNP) analysis
Summary
A Nanopore Detector provides a means to transduce single molecule events into observable channel current changes. The nanopore-based detector can directly measure molecular characteristics in terms of the blockade properties of individual molecules – this is possible due to the kinetic information that is embedded in the blockade measurements, where the adsorptiondesorption history of the molecule to the surrounding channel, and the configurational changes in the molecule itself, imprint on the ionic flow through the channel This rich source of information offers prospects for DNA sequencing and single nucleotide polymorphism (SNP) analysis. A New Method for Single Molecule Detection and Characterization Angstrom precision structures for numerous DNA, RNA, and protein molecules have been revealed by X-ray diffraction analysis and NMR spectroscopy These approaches rely upon average properties of very large numbers of molecules and are often biased towards crystallization and NMR conformer structures different from those present in solution under any conditions, physiological conditions in particular. The nanopore detector presents the possibility of observing conformational change within a molecule (see [26] for latest results), something not addressed by other methods
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