Nanopore Detection of Nucleic Acid Displacement Circuits
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Solution Overview
Problem
Current DNA circuit detection methods, relying on fluorescence reporters, are limited by spectral overlap, restricting the number of unique signals that can be detected in a single-pot reaction, and are not scalable for complex DNA circuits.
Innovation Solution
A nanopore system is used to detect nucleic acid strand displacement circuits by translocating displaced output strands through a nanopore, measuring ion currents, and associating these currents with input strands, enabling scalable and multiplexed detection using machine learning models to differentiate unique barcode sequences.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If fluorescence reporters are used to detect DNA circuits, then sensitivity and real-time reporting are improved, but the number of unique signals that can be detected in parallel is limited due to spectral overlap
Solution Approach 1:
The patent replaces the optical detection system (fluorescence reporters and spectrofluorometers) with an electrical detection system using nanopores. Each nanopore measures ionic current blockades as DNA strands translocate through it, converting optical signal detection into electrical signal detection. This substitution eliminates spectral overlap limitations while maintaining single-molecule sensitivity and real-time detection capabilities.
Solution Approach 2:
The patent changes the detection parameter from optical properties (fluorescence emission wavelengths) to electrical properties (ionic current blockades). By measuring changes in ionic current rather than light emission, the system can distinguish between multiple different DNA sequences simultaneously without the spectral overlap that limits fluorescence-based methods. Each DNA strand produces a unique current signature based on its sequence and translocation kinetics.
2Quantity of substance
If multiple fluorescence reporters are used to increase the number of detectable signals, then the number of unique signals increases, but spectral overlap increases and detection precision decreases
Solution Approach 1:
The patent replaces the optical detection system (fluorescence reporters and spectrofluorometers) with an electrical detection system using nanopores. Each nanopore measures ionic current blockades as DNA strands translocate through it, converting optical signal detection into electrical signal detection. This substitution eliminates spectral overlap limitations while maintaining single-molecule sensitivity and real-time detection capabilities.
Solution Approach 2:
The patent changes the detection parameter from optical properties (fluorescence emission wavelengths) to electrical properties (ionic current blockades). By measuring changes in ionic current rather than light emission, the system can distinguish between multiple different DNA sequences simultaneously without the spectral overlap that limits fluorescence-based methods. Each DNA strand produces a unique current signature based on its sequence and translocation kinetics.
3Adaptability or versatility
If DNA circuit complexity increases, then the information processing capability improves, but the scalability of detection methods becomes insufficient
Solution Approach 1:
The patent creates a universal nanopore detection platform that can read out any DNA circuit output regardless of sequence. The nanopore system performs multiple functions: it detects single-molecule translocation events, distinguishes different DNA sequences through current signatures, quantifies strand displacement kinetics, and scales to multiplexed detection. This universal platform can handle increasingly complex DNA circuits without requiring detection method modifications.
Solution Approach 2:
The nanopore detection system requires minimal intervention and preparation. DNA strands are loaded into the nanopore system and automatically translocate through the pore under an applied voltage, generating detectable current signals without requiring enzymatic amplification, optical labeling, or complex sample preparation. The system self-detects and records the translocation events, enabling scalable detection of complex DNA circuit outputs.
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 allows for real-time, multiplexed detection and quantification of nucleic acid strand displacement reactions, overcoming the limitations of fluorescence-based methods by enabling the differentiation of multiple barcode sequences and scaling to larger barcode sets, thus enhancing the readout capabilities of DNA circuits.
Implementation Method 1
measuring an ion current through the nanopore when the displaced output strand is in the tunnel to provide a current pattern corresponding to a portion of the displaced output strand
Implementation Method 2
translocating the displaced output strand through the nanopore from the first conductive liquid medium towards the second conductive liquid medium
Implementation Method 3
permitting hybridization of the input strand to the partner strand along a second portion of the partner strand that partially overlaps with the first portion of the partner strand, thereby displacing the output strand from the double stranded complex
Data Source
AI summary
The disclosure provides compositions, systems, and related method for using nanopore-based detection of nucleic acid displacement circuits. In some embodiments, output strands contain orthogonal barcode sequences that are captured by nanopore systems to produce a unique and recognizable current signal. Machine learning models can be used to differentiate a plurality of current signals and, therefore, monitor detection and quantification of multiple nucleic acid displacement circuits in a single pot reaction.