UMI Adaptor Tagging for Rare cfDNA Copy Number Detection
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Solution Overview
Problem
Existing methods for genetic analysis, particularly in detecting rare copy number changes in biological samples, lack the sensitivity to identify mutations at very minor allele frequencies, especially in cell-free DNA (cfDNA) samples.
Innovation Solution
The use of adaptors with unique molecular identifiers (UMIs) and sample tags to generate genomic DNA libraries, allowing for high-resolution detection of genetic changes by forming complexes with capture probes, followed by primer extension, amplification, and sequencing to identify genetic variations such as copy number changes.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If conventional SNP genotyping methods are used, then allele frequencies can be detected at common levels (~100%, 50%, or 0%), but the methods lack sensitivity to detect rare mutations at very minor allele frequencies in cfDNA samples
Solution Approach 1:
The adaptor is segmented into distinct functional regions: a UMI region for molecular identification, a sample index region for sample identification, and an amplification region. This segmentation allows each region to perform its specific function optimally, with the UMI region enabling precise tracking of individual DNA molecules to detect rare mutations even in small cfDNA samples
Solution Approach 2:
The UMI is incorporated into the adaptor during the initial library preparation step, before amplification and sequencing. This preliminary tagging of each DNA molecule with a unique identifier allows for accurate reconstruction of the original DNA population and detection of rare mutations that would otherwise be lost in the noise of amplification and sequencing variability
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
The method achieves sensitive detection of copy number variations in a small fraction of total DNA, enabling accurate identification of genetic changes indicative of disease states, even at minor allele frequencies.
Implementation Method 1
adaptors with unique molecular identifiers (UMIs) and sample tags to generate genomic DNA libraries, allowing for high-resolution detection of genetic changes by forming complexes with capture probes
Implementation Method 2
contacting the genomic DNA library with a plurality of capture probes that specifically bind to a DNA target region, thereby forming complexes between the capture probes and DNA library fragments comprising the DNA target region
Implementation Method 3
performing primer extension and amplification of the DNA library fragments comprising the DNA target region
Implementation Method 4
amplification of the DNA library fragments comprising the DNA target region
Data Source
Figure 1~2
Figure 3
Figure 4A~4B
AI summary
The present invention includes compositions and methods useful for the detection of a mutational change, SNP, translocation, inversion, deletion, change in copy number, or other genetic variation within a sample of cellular genomic DNA or cell-free DNA (cfDNA). In some embodiments, the compositions and methods of the present invention provide an extremely high level of resolution that is particularly useful in detecting copy number variations in a small fraction of the total cfDNA from a biological sample (e.g., blood).