Non-human nucleic acid controls for human DNA assay validation
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Solution Overview
Problem
Current DNA processing and testing methods for detecting mutations and methylation status in human samples require specific control nucleic acids that do not cross-react with human DNA, posing challenges in assay validation and error detection.
Innovation Solution
The use of non-human nucleic acids, such as zebrafish DNA with specific methylation patterns, that undergo normal testing and processing to serve as run controls, providing a normal range of results and avoiding cross-reactivity with human nucleic acid detection assays.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If human DNA control molecules are used for assay validation, then the control can be measured post-extraction to assess efficiency, but the control may cross-react with detection assays designed for human nucleic acids
Solution Approach 1:
The patent uses non-human nucleic acids (from species such as zebrafish, honey bee, or other non-mammalian organisms) as intermediary control molecules. These intermediaries serve the function of process controls while being sufficiently distinct from human DNA sequences to avoid cross-reactivity with human-specific detection assays, thus resolving the contradiction between needing reliable controls and avoiding harmful cross-reactivity
Solution Approach 2:
The patent changes the species origin parameter of the control nucleic acid from human to non-human organisms. This parameter change maintains the functional properties needed for process control (such as methylation patterns and DNA structure) while altering the sequence composition to eliminate cross-reactivity with human DNA detection assays
2Object-affected harmful factors
If non-human nucleic acids are used as process controls, then cross-reactivity with human DNA assays is avoided, but the control must still resemble human nucleic acid properties to be effective
Solution Approach 1:
The patent applies local quality by selecting specific regions or genes from non-human organisms that possess local properties similar to human DNA (such as methylation patterns at CpG islands, GC content, and DNA secondary structure) while maintaining global sequence differences. This allows the control to behave like human DNA during extraction and processing steps while remaining distinguishable and non-cross-reactive in detection assays
Data Source
AI summary
The present invention provides synthetic DNA strands that find use as process controls in DNA processing and nucleic acid testing methods. In particular, provided herein are synthetic methylated DNA strands of known composition for use as control molecules in DNA testing, e.g., of mutations and/or methylation of DNA isolated from non-fish samples, such as human samples.