Adapter QC Nucleosides for Accurate DNA Modification Detection
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Solution Overview
Problem
Existing single-nucleotide resolving assays for detecting epigenetic variants or nucleoside modifications suffer from false positive and false negative signals due to incomplete or erroneous conversion of nucleosides, particularly in methods like bisulfite and TAPS, leading to inaccurate methylation assessments.
Innovation Solution
Incorporating quality control nucleosides in adapters that maintain or alter base pairing specificity differently from the DNA sample nucleosides, allowing for the detection of suboptimal conversion and subsequent correction of false signals through sequence analysis and weighting.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If conversion procedure is applied to detect modified nucleosides, then detection capability is improved, but false positive and false negative signals increase due to incomplete or erroneous conversion
Solution Approach 1:
The patent introduces an intermediary conversion step that transforms modified nucleosides into detectable forms with altered base-pairing specificity. This intermediary conversion allows indirect detection of modifications while enabling quality control measures to assess conversion completeness and correct false signals through control nucleotides and computational methods
Solution Approach 2:
The patent implements feedback mechanisms by incorporating control nucleotides in adapters that undergo the same conversion procedure as sample nucleotides. The conversion status of control nucleotides provides feedback on conversion efficiency, enabling identification and correction of false positive and false negative signals in the methylation detection results
2Reliability
If quality control nucleosides are incorporated in adapters, then false signal detection is improved, but adapter sequence complexity increases
Solution Approach 1:
The patent segments the adapter sequence into distinct functional regions: control nucleotide regions and sample binding regions. This segmentation allows the control nucleotides to independently monitor conversion status without interfering with sample nucleotide detection, managing complexity through functional separation
Solution Approach 2:
The adapters serve multiple functions simultaneously: they facilitate sample nucleotide binding, undergo conversion procedure, provide control nucleotides for quality monitoring, and enable sequencing. This multi-functionality reduces the need for separate control mechanisms, managing complexity through integrated design
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
Enhances the accuracy of methylation detection by identifying and correcting false positives and negatives, ensuring reliable interpretation of methylation data.
Implementation Method 1
the conversion procedure is selected to change the base pairing specificity of quality control nucleosides in the adapters
Implementation Method 2
change their base-pairing specificity
Data Source
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AI summary
Provided herein are methods for monitoring false negative and/or false positive detection of modified nucleosides in DNA in a sample using a base-pairing conversion procedure. The methods use nucleosides having known nucleoside identity and known modification status in adapters ligated to the DNA. In certain aspects, the disclosure relates to methods for improving the quality control of such methods.