Enzyme Translocators in Nanogap for Single Base Resolution Sequencing
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Solution Overview
Problem
Current nucleic acid sequencing technologies face challenges in achieving single base pair resolution with long reads, requiring either reconstruction from small reads or repeated runs, and struggle to combine scalability and speed with high accuracy.
Innovation Solution
The method employs an electrochemical nanoelectrode sensor with a dielectric layer between electrodes, where a polymerase enzyme is attached to target a polynucleotide strand. Modified nucleotides with electroactive labels covalently bound via an ester group are used, allowing for single base resolution sequencing by inducing electron flow and producing measurable electrical signals.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If semiconductor-based electrical detection is used, then scalability and speed are improved, but measurement precision deteriorates
Solution Approach 1:
The patent introduces an enzyme translocator as an intermediary component that binds to the polynucleotide strand and facilitates its translocation through the nanogap. This enzyme mediator enables precise single-base-step movement, achieving high measurement precision while maintaining scalability through parallel processing of multiple strands simultaneously.
Solution Approach 2:
The patent implements a nested structure where the polynucleotide strand is threaded through a nanogap, the enzyme translocator is positioned within the gap to interact with the strand, and electrodes are arranged to detect signals at the nanoscale. This nested configuration enables precise spatial control for single base pair resolution while allowing high-throughput parallel sequencing.
2Measurement precision
If sequencing by synthesis technologies are used, then measurement precision is improved, but device complexity increases
Solution Approach 1:
The patent extracts and isolates the critical sequencing function to a minimal nanoscale platform: a simple nanogap between two electrodes with an enzyme translocator. This extracted core mechanism achieves high precision without the complex optical systems, multiple reagent channels, and sophisticated instrumentation required by traditional sequencing by synthesis technologies.
3Length of moving object
If long reads are achieved through reconstruction from small reads, then read length is improved, but loss of information increases
Solution Approach 1:
The patent enables continuous translocation of the polynucleotide strand through the nanogap in single-base steps, allowing uninterrupted sequential detection of nucleotides over long distances. This continuous action produces long reads with high fidelity without requiring fragmented assembly from multiple short reads, thereby minimizing information loss.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
This approach enables long reads with single base pair resolution, overcoming the limitations of existing methods by providing a scalable and accurate sequencing technique that combines the benefits of semiconductor-based electrical detection with the high accuracy of sequencing by synthesis technologies.
Implementation Method 1
applying a first potential to the first electrode and a second potential to the second electrode to induce electron flow between the first and second electrodes to produce a measurable electrical signal when an electroactive label is present in the sensing zone
Data Source
AI summary
A method for nucleic acid sequencing includes providing at least one device comprising a first electrode and a second electrode separated by a dielectric layer, and a polymerase enzyme attached to the surface of the dielectric layer. The dielectric layer induces an electroactive molecule to interact with the electrodes to complete an electrical circuit. The polymerase enzyme targets a polynucleotide strand to the dielectric layer. A sample including a polynucleotide strand and modified nucleotides having an electroactive label covalently bound to the 3′—OH of a sugar ring of the nucleotide via an ester group is provided to the at least one device. Potentials are applied to each electrode to induce electron flow between the electrodes to produce a measurable electrical signal when an electroactive label is present in the dielectric layer. Electrical signals from the electrodes are detected to determine when a modified nucleotide is present in the dielectric layer.