Linked DNA Fragment Sequencing for Direct Error Detection
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Solution Overview
Problem
High-throughput genomic sequencing platforms suffer from inaccuracies due to error propagation in amplification and sequencing processes, leading to misalignment and misidentification of mutations, which current error detection strategies are costly and inefficient.
Innovation Solution
Linking two identical nucleic acid fragments together to form a complex, allowing errors to be identified during sequencing by comparing the agreement or disagreement between the fragments, thereby reducing error rates and sequencing costs.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If standard barcode sequencing methods use tens to hundreds of copies of the same template to catch errors, then error detection capability is improved, but sequencing costs increase and sequencing bandwidth is consumed
Solution Approach 1:
The patent combines multiple templates into a single linked cluster structure, where multiple templates are physically connected through adapter sequences. This allows error detection across multiple templates without requiring separate clusters for each template, thereby reducing the total number of clusters needed while maintaining error detection capability.
Solution Approach 2:
The linked cluster structure serves multiple functions simultaneously: it amplifies multiple templates, enables error detection through comparison, and reduces sequencing bandwidth consumption. The single cluster design provides both amplification and error detection capabilities that would otherwise require separate systems.
2Productivity
If only one fragment seeds a cluster in traditional amplification methods, then amplification efficiency is improved, but error propagation occurs throughout the entire cluster
Solution Approach 1:
Multiple templates are merged into a single linked cluster through adapter sequences, allowing them to be amplified together in one cluster rather than requiring separate clusters. This maintains amplification efficiency while enabling cross-template error detection through the linking structure.
Solution Approach 2:
The linked cluster structure provides built-in feedback for error detection by maintaining physical connections between multiple templates. During sequencing, discrepancies between linked templates can be detected and used to identify amplification errors, providing real-time error correction capability.
3Measurement precision
If tens to hundreds of clusters are used to create a sample pool for error comparison, then error determination accuracy is improved, but sequencing bandwidth is consumed and costs increase
Solution Approach 1:
Multiple templates that would traditionally require separate clusters are merged into single linked clusters. This consolidation maintains the ability to compare multiple templates for error determination while using fewer total clusters, thereby preserving sequencing throughput and reducing costs.
Solution Approach 2:
The patent creates multiple copies of templates within a single linked cluster structure rather than requiring multiple separate clusters. This copying approach maintains sufficient template diversity for error detection while consolidating the physical infrastructure needed for sequencing.
Data Source
AI summary
The invention generally relates to sequencing library preparation methods. In certain embodiments, two template nucleic acids are joined together by a linking molecule, such as a PEG derivative. The linked template nucleic acids is amplified, creating linked amplicons.