Cell-Free DNA Mutation Detection via Molecular Barcoding
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Solution Overview
Problem
Current methods for detecting genetic diseases, particularly cancer, using cell-free DNA are limited by the need for improved techniques to accurately identify copy number variations and rare mutations in bodily fluids, which are essential for early detection and monitoring.
Innovation Solution
A method involving sequencing of extracellular polynucleotides from bodily samples, using unique barcodes for identification, filtering, mapping, and normalization to detect copy number variations and rare mutations by comparing sequence reads to reference sequences, and generating genetic profiles for characterization.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If traditional sequencing methods are used to detect genetic aberrations in cell-free DNA, then the detection process becomes feasible, but the sensitivity and accuracy for detecting rare mutations and copy number variations are insufficient
Solution Approach 1:
The patent segments the detection process into multiple independent steps: (1) attaching unique molecular barcodes to individual cell-free DNA molecules before amplification, (2) performing massively parallel amplification of barcoded molecules, (3) sequencing the amplified products, and (4) using computational algorithms to trace reads back to parent molecules and detect aberrations. This segmentation allows each step to be optimized independently, achieving both high sensitivity for rare mutations and high accuracy for copy number variation detection.
Solution Approach 2:
The patent introduces molecular barcodes as an intermediary element that bridges the gap between low-abundance cell-free DNA molecules and the detection capability of sequencing instruments. These barcodes serve as unique identifiers that allow multiple copies of the same original molecule to be tracked and counted, enabling detection of rare mutations and quantitative analysis of copy number variations with high sensitivity and precision.
2Loss of time
If cell-free DNA from bodily fluids is analyzed for early disease detection, then early diagnosis becomes possible, but the low concentration of disease-associated DNA fragments makes detection difficult
Solution Approach 1:
The patent performs preliminary enrichment and barcoding of cell-free DNA molecules before amplification and sequencing. By attaching unique molecular barcodes to individual DNA molecules at the earliest stage, the method enables subsequent massively parallel amplification that preserves the original molecular information. This preliminary action allows the system to work effectively with the low concentrations of disease-associated DNA fragments present in bodily fluids, enabling early detection without requiring high DNA concentrations.
3Reliability
If massively parallel sequencing is performed to increase detection sensitivity, then more genetic aberrations can be detected, but the complexity of data analysis and computational requirements increase
Solution Approach 1:
The patent uses molecular barcodes as copies of the original DNA molecule identity that are preserved through amplification. Each barcode serves as a simplified representation of the parent molecule, allowing computational algorithms to efficiently trace thousands of sequencing reads back to their parent molecules and accurately count molecular populations. This copying approach reduces data processing complexity by providing direct molecular identifiers rather than requiring complex de novo assembly or analysis of raw sequencing data.
Data Source
AI summary
The present disclosure provides a system and method for the detection of rare mutations and copy number variations in cell free polynucleotides. Generally, the systems and methods comprise sample preparation, or the extraction and isolation of cell free polynucleotide sequences from a bodily fluid; subsequent sequencing of cell free polynucleotides by techniques known in the art; and application of bioinformatics tools to detect rare mutations and copy number variations as compared to a reference. The systems and methods also may contain a database or collection of different rare mutations or copy number variation profiles of different diseases, to be used as additional references in aiding detection of rare mutations, copy number variation profiling or general genetic profiling of a disease.