Unique Molecular Barcodes for Cell-Free DNA Sequencing
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Solution Overview
Problem
Current methods for analyzing cancer-derived cell-free DNA face challenges due to low nucleic acid yields and sequencing errors, limiting the sensitivity and accuracy of mutation profiling.
Innovation Solution
The use of a pool of unique adaptors with specific barcode structures for nucleic acid analysis, including double-stranded and single-stranded barcodes, to enhance sequencing accuracy and sensitivity by reducing errors and identifying consistent mutations.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If conventional sequencing methods are used to analyze cell-free DNA, then the analysis can be performed with standard techniques, but sequencing errors limit analytical sensitivity and mutation profiling accuracy
Solution Approach 1:
The invention segments the DNA analysis process by attaching unique molecular barcodes to individual DNA molecules before amplification. This allows tracking of original molecules through multiple PCR cycles, enabling error correction by comparing consensus sequences from multiple amplifications of the same original molecule, thereby improving analytical sensitivity while maintaining reliability
Solution Approach 2:
The invention introduces molecular barcodes as an intermediary element between the original DNA and the sequencing process. These barcodes serve as unique identifiers that allow differentiation of original DNA molecules from amplification artifacts, enabling accurate mutation profiling by distinguishing true genetic variations from sequencing errors
2Quantity of substance
If standard PCR amplification is used to increase nucleic acid yield, then sufficient material for sequencing is obtained, but artifacts are introduced that reduce analysis accuracy
Solution Approach 1:
The invention performs preliminary action by attaching unique molecular barcodes to DNA molecules before amplification. This pre-tagging allows subsequent tracking and verification of amplification fidelity, enabling identification and correction of PCR-introduced artifacts while maintaining the necessary nucleic acid yield for sequencing
3Productivity
If high-throughput sequencing is performed on low-abundance tumor DNA, then detection capability is enhanced, but the low nucleic acid yields from tumor shedding limit the effectiveness of current methods
Solution Approach 1:
The invention uses molecular barcodes as copies that can be replicated alongside the original DNA through PCR amplification. Each barcode serves as a unique identifier that is copied faithfully (with trackable fidelity) through multiple amplification cycles, enabling high-throughput sequencing of low-abundance tumor DNA by creating sufficient copies for detection while maintaining the ability to trace back to original molecules for error correction
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 significantly improves the sensitivity of detecting tumor-derived nucleic acids, allowing for more accurate mutation profiling and monitoring of cancer, with reduced error rates and improved detection limits.
Implementation Method 1
each adaptor comprises a double stranded portion at a proximal end and two single stranded portions at a distal end, wherein the double stranded portion comprises a double-stranded barcode
Implementation Method 2
attaching a pool of adaptors according to claims 1-6 to both ends of a plurality of double-stranded nucleic acids via the double stranded portions of the adaptors
Data Source
Figure 1a~1c
Figure 1d
Figure 1e
AI summary
Disclosed herein are polynucleotide adaptors and methods of use thereof for identifying and analyzing nucleic acids, including cell-free nucleic acids from a patient sample. Also disclosed herein are methods of using the adaptors to detect, diagnose, or determine prognosis of cancers.