Sequencing by Emergence for Long-Range Haplotype Resolution
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Solution Overview
Problem
Current nucleic acid sequencing technologies face challenges with high costs and inefficiencies due to reliance on reagents and time, limited read lengths, and the need for reference genomes, which hinder the efficient sequencing of large genomes like the human genome.
Innovation Solution
A method involving the transient and reversible binding of oligonucleotide probes to fixed, elongated nucleic acids on a substrate, allowing for optical activity measurement and sequence determination through imaging, enabling efficient sequencing without the need for extensive reagent exchange or amplification.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If gel electrophoresis-based sequencing methods (Sanger or chemical degradation) are used, then sequencing accuracy is achieved, but the process becomes time-consuming and expensive
Solution Approach 1:
The patent replaces the mechanical gel electrophoresis separation system with a direct imaging system that detects fluorescently labeled nucleotides as they are incorporated into growing DNA strands. This substitution eliminates the time-consuming gel running and analysis steps while maintaining sequencing accuracy through optical detection of each incorporated nucleotide.
Solution Approach 2:
The sequencing process operates continuously by maintaining DNA polymerase activity and repeatedly adding fluorescently labeled nucleotides to growing strands without interruption. The system continuously images the incorporation events, allowing multiple bases to be sequenced in a single continuous reaction rather than through discrete, time-consuming steps.
2Measurement precision
If gel electrophoresis-based sequencing methods are used, then sequencing accuracy is achieved, but the cost increases significantly
Solution Approach 1:
The patent replaces the expensive gel electrophoresis infrastructure with a more cost-effective direct imaging system. By using standard fluorescent microscopy or similar optical systems to detect nucleotide incorporation, the method eliminates the need for specialized gel apparatus, reagents, and labor-intensive processing while maintaining accurate base calling through optical signal detection.
3Productivity
If Illumina sequencing with reversible terminators is used, then large-scale sequencing is achieved, but substantial upfront sample processing and clonal amplification are required
Solution Approach 1:
The patent extracts and eliminates the complex clonal amplification and library preparation steps that are central to Illumina sequencing. By using single-molecule sequencing with direct detection of nucleotide incorporation, the method processes native DNA samples without requiring conversion to amplified clusters, thereby reducing sample processing complexity while maintaining high throughput through parallel imaging of multiple molecules.
4Length of stationary object
If read length is increased in commercial sequencing systems, then more reagent supply cycles are needed, but this increases both time and cost
Solution Approach 1:
The patent maintains continuous sequencing operation by keeping DNA polymerase active throughout the process and repeatedly adding fluorescently labeled nucleotides without requiring reagent exchange or system interruption. This continuous operation allows read lengths to extend to thousands of bases without the time penalties associated with repeated reagent loading cycles, as the imaging system continuously captures incorporation events throughout the extended reaction.
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 accurate, long-range haplotype-resolved sequencing with reduced reagent consumption and time, improving the efficiency and cost-effectiveness of nucleic acid sequencing while maintaining high accuracy.
Implementation Method 1
binding molecular probes to one or more units of a double stranded target molecule
Implementation Method 2
The detection methods used in the most evolved form of Sanger sequencing and the currently dominant Illumina technology involve fluorescence
Data Source
AI summary
The invention is a method of sequencing polymers in which the sequence of one or more polymers is determined through an emergent property of the binding interactions of a repertoire of molecular probes to the polymer(s).