Double CsgG Pore Nanopore for Nucleotide Discrimination
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Solution Overview
Problem
Current nanopore sensing technologies face challenges in accurately controlling nucleic acid movement through pores and discriminating between nucleotides, particularly for polynucleotides with homopolymeric stretches.
Innovation Solution
The use of double pores composed of two CsgG pores in series, along with novel monomers that enhance interactions between the pores, facilitate interaction with analytes, and increase the length of the pore constriction, to improve analyte detection and characterization.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If a single CsgG pore is used for nanopore sensing, then the device structure is simple, but the accuracy of nucleotide discrimination and control of nucleic acid movement is insufficient
Solution Approach 1:
The single pore is divided into two separate CsgG pores arranged in series, creating a double pore structure. Each pore acts as an independent sensing unit with its own reader head, allowing for more accurate nucleotide discrimination while maintaining the modular nature of the original single pore design
Solution Approach 2:
Two CsgG pores are merged into a single functional unit where their constrictions are positioned in series along the transmembrane axis. This combination allows the double pore to function as an integrated sensing system that improves measurement precision while managing the increased structural complexity through coordinated design
2Loss of information
If the reader head is made short and sharp, then the device complexity is low, but additional information for accurate base calling is limited
Solution Approach 1:
The reader head is extended along the transmembrane axis (adding length in one dimension) to include additional amino acid residues that provide extra information for base calling. This dimensional extension allows more nucleotides to be sensed simultaneously without fundamentally changing the pore's operational principles
3Measurement precision
If mutations are made to sharpen the reader head, then nucleotide discrimination is improved, but the ability to handle homopolymeric stretches is insufficient
Solution Approach 1:
The sensing function is segmented into two separate pores, each with its own reader head. This segmentation allows the system to process homopolymeric stretches more effectively by distributing the sensing load across multiple reader heads, providing redundant information for accurate base calling in repetitive sequences
Solution Approach 2:
The reader head parameters are modified by extending its length and changing amino acid compositions. These parameter changes enhance the system's adaptability to different sequence types, including homopolymeric stretches, while maintaining the discrimination capability provided by the constricted geometry
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
The double pore configuration enhances the accuracy of nucleotide discrimination and improves the characterization of polynucleotides, including those with homopolymeric stretches, by providing additional information and longer reader heads.
Implementation Method 1
measuring voltage-driven ionic transport through the pore in the presence of analyte molecules
Data Source
AI summary
Provided is a method of characterising a polynucleotide using a transmembrane pore, wherein the pore is a double pore comprising a first Csg G pore, or a homologue thereof, and second CsgG pore, or a homologue thereof.