Linked Target Capture for Consensus-Based Error Detection
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Solution Overview
Problem
High-throughput genomic sequencing platforms suffer from inaccuracies due to amplification and sequencing errors, leading to sequence misalignment and misidentification of mutations, with existing error detection methods being costly and inefficient.
Innovation Solution
The method involves linking two or more nucleic acid fragments, such as both strands of a duplex DNA molecule, to increase information density and reduce error rates by comparing bases across strands, using linked primers and probes to capture and amplify these fragments, and incorporating them into sequencing clusters without unique tagging.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If standard barcode sequencing methods use tens to hundreds of copies of the same template or ten to hundreds of clusters to create a sample pool for comparison, then error detection capability is improved, but sequencing cost and bandwidth consumption increase
Solution Approach 1:
The patent merges multiple sequencing reads that originate from the same template molecule into a single consensus sequence. By linking multiple reads through template origin tracking, the method combines information from multiple copies without requiring them to be sequenced separately, thereby reducing the total number of clusters needed while maintaining error detection capability.
Solution Approach 2:
The patent creates a consensus sequence as a copy that represents the true template sequence by comparing multiple reads. This consensus copy eliminates errors present in individual reads, providing accurate sequence information without needing to sequence numerous separate clusters.
2Productivity
If a single fragment seeds a cluster in amplification, then amplification efficiency is improved, but error propagation increases
Solution Approach 1:
The patent implements feedback by comparing multiple reads that originate from the same template. The consensus sequence generation process uses feedback from multiple independent reads to identify and correct errors, allowing the system to maintain high amplification efficiency while eliminating error propagation through comparative analysis.
Solution Approach 2:
The patent segments the sequencing process into multiple independent reads from the same template, then analyzes each read separately before combining them into a consensus. This segmentation allows errors in individual reads to be identified and corrected while maintaining the efficiency of single-fragment seeding.
3Measurement precision
If base calling and alignment algorithms are used to improve accuracy, then sequence alignment quality is improved, but the impact of amplification and sequencing errors on quality remains
Solution Approach 1:
The patent applies preliminary anti-action by generating a consensus sequence before alignment and base calling processes. The consensus sequence generation preemptively corrects amplification and sequencing errors by comparing multiple reads, so that subsequent alignment and base calling operate on already-corrected data, eliminating the harmful effects of errors at their source.
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 enhances base calling accuracy by differentiating between true variants and errors introduced during amplification or sequencing, reducing sequencing costs and improving sequencing throughput.
Implementation Method 1
the fragment is amplified and the amplification products attach to the solid support proximate to the seeding fragment
Implementation Method 2
DNA fragments are attached to a solid support, such as a channel wall
Data Source
AI summary
The invention generally relates to sequencing library preparation methods. In certain embodiments, two or more template nucleic acids are joined together by a linking molecule, such as a PEG derivative. Identical copies of a nucleic acid fragment or both strands of a duplex fragment may be linked together. The linked nucleic acids are amplified, creating linked amplicons. Emulsion PCR with linked primers creates linked template nucleic acids for seeding sequencing clusters and errors can be readily identified by their presence on only one of the linked fragments.