Cell-free DNA Methylation Enrichment via Differential Partitioning
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Solution Overview
Problem
Current methods for analyzing cell-free DNA in liquid biopsies face challenges in accurately detecting nucleobase modifications due to low concentration and heterogeneity, requiring improved techniques for isolating and processing fractions for effective analysis.
Innovation Solution
A method involving subjecting DNA to a procedure that differentially affects different nucleobases, followed by partitioning into subsamples based on nucleobase modifications and sequencing to distinguish between them, providing combined information on methylation levels and specific modifications.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Ease of operation
If liquid biopsy is used to detect cancer, then noninvasive detection is achieved, but detection accuracy is reduced due to low concentration and heterogeneity of cell-free DNA
Solution Approach 1:
The patent segments cell-free DNA molecules into different partitions based on their methylation characteristics. By treating the DNA sample to differentially affect methylated and unmethylated nucleobases, then partitioning into multiple subsamples, the method enables accurate detection of methylation patterns despite the low concentration and heterogeneity of cell-free DNA in liquid biopsy
Solution Approach 2:
The patent introduces an intermediary procedure that differentially affects methylated versus unmethylated nucleobases before partitioning. This intermediary treatment step enables the subsequent partitioning process to effectively separate and enrich methylated DNA fragments, thereby improving detection accuracy while maintaining the noninvasive nature of liquid biopsy
2Loss of information
If DNA is partitioned into multiple subsamples based on methylation, then detailed epigenetic information is obtained, but process complexity increases
Solution Approach 1:
The patent divides the DNA sample into multiple partitions based on methylation characteristics, with each partition containing DNA molecules with similar methylation patterns. This segmentation approach preserves detailed epigenetic information by maintaining the distribution of different methylation levels across partitions, while the systematic nature of the partitioning process manages complexity through standardized procedures
Solution Approach 2:
The patent changes the chemical state of nucleobases through a treatment procedure that differentially affects methylated versus unmethylated bases. This parameter change enables subsequent partitioning based on methylation status, preserving epigenetic information while the controlled nature of the chemical transformation keeps the process manageable
3Measurement precision
If bisulfite conversion is used to detect methylation, then methylation status is identified, but DNA damage and loss occur
Solution Approach 1:
The patent extracts and enriches methylated DNA fragments through partitioning before subjecting them to bisulfite conversion. By taking out and concentrating the methylated DNA of interest into specific partitions, the method improves methylation detection accuracy while reducing the overall amount of DNA subjected to damaging bisulfite treatment, thereby preserving DNA integrity
Solution Approach 2:
The patent performs partitioning and enrichment of methylated DNA before applying bisulfite conversion. This preliminary action concentrates the target methylated DNA while minimizing the exposure of all DNA to the harsh bisulfite treatment, thereby maintaining better DNA integrity while still achieving accurate methylation detection
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 the analysis of cell-free DNA by providing more detailed epigenetic information, improving the detection of cancer indicators through better differentiation of methylation patterns and modifications.
Implementation Method 1
subjecting the population of nucleic acids to a procedure that affects a first nucleobase in the nucleic acids differently from a second nucleobase in the nucleic acids
Implementation Method 2
partitioning the population of nucleic acids into a plurality of partitions by contacting the nucleic acids with a methyl-binding reagent
Data Source
AI summary
Provided herein is a DNA analysis method comprising a procedure that affects a first nucleobase in the DNA differently from a second nucleobase in the DNA of the first subsample; partitioning a sample into at least a first subsample and a second subsample, wherein the first subsample comprises DNA (e.g., cell-free DNA) with a nucleobase modification in a different proportion than the second subsample; and DNA is sequenced to distinguish the first nucleobase from the second nucleobase. Also provided is a combination comprising first and second populations of captured DNA, wherein the first population comprises or was derived from DNA with a nucleobase modification in a different proportion than the second population, and wherein the first population comprises a form of a first nucleobase originally present in the DNA with altered base pairing specificity and a second nucleobase without altered base pairing specificity.