Differential k-Mer Amplification for Pathogen DNA Enrichment
Find Innovative SolutionsGenerate Solutions
Solution Overview
Problem
Current methods for enriching nucleic acid sequences of microorganisms and viruses in complex samples, such as blood, are inefficient and costly, particularly when the host and target organisms have similar genomic features, leading to high background levels of human DNA that hinder effective sequencing.
Innovation Solution
The use of differentially abundant k-mers, specifically six-nucleotide sequences, to selectively amplify pathogen DNA by identifying k-mers that show differences in frequency and/or context between the target microorganism and human genomes, allowing for preferential amplification using techniques like multiple displacement amplification (MDA).
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Quantity of substance
If conventional host DNA removal techniques are used, then human DNA depletion is achieved, but the process becomes costly and requires additional sample preparation steps
Solution Approach 1:
The method performs preliminary identification of organism-specific k-mers before the actual sequencing process. By pre-determining which k-mers are unique to the host versus pathogen, the system prepares the discrimination criteria in advance, eliminating the need for complex post-extraction separation steps and reducing overall workflow complexity
Solution Approach 2:
The invention extracts and utilizes only the discriminatory k-mer sequences from the genomic data. Instead of physically removing host DNA through complex depletion techniques, the method extracts the informational signature (k-mers) that enables computational distinction between host and pathogen sequences during analysis
2Quantity of substance
If host DNA depletion techniques are applied, then background human DNA is reduced, but the cost increases
Solution Approach 1:
The method uses short, inexpensive k-mer sequences (typically 6-12 nucleotides long) as discriminative markers. These short k-mers are computationally generated and can be rapidly processed, replacing costly physical depletion reagents and procedures with low-cost computational analysis
3Measurement precision
If conventional enrichment techniques are used, then pathogen DNA can be detected, but the method fails when host and target organisms have similar genomic features
Solution Approach 1:
The method identifies and utilizes local unique characteristics (specific k-mer sequences) within the pathogen genome that differ from the host genome. By focusing on these locally unique regions rather than requiring global genomic differences, the system can distinguish between organisms even when they share many overall genomic features
Solution Approach 2:
The approach changes the parameter of discrimination from broad genomic categories to specific k-mer sequence frequencies and compositions. By analyzing the presence, absence, or frequency of specific short sequences rather than overall genomic similarity, the system achieves discrimination capability even between closely related organisms
4Measurement precision
If direct sequencing of complex biological samples is performed, then pathogen detection is attempted, but high background levels of host DNA hamper effective sequencing
Solution Approach 1:
The k-mer sequences serve as an intermediary between the raw sequencing data and the final pathogen detection result. By introducing this computational layer that counts and compares k-mer frequencies against reference databases, the system effectively filters out host background without requiring physical removal, enabling sensitive pathogen detection even in high-host-DNA samples
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 enrichment of microorganism and virus nucleic acid sequences, reducing background noise and improving sequencing efficiency without additional sample preparation steps, thereby making the process more cost-effective and effective.
Implementation Method 1
amplifying the pathogen DNA sequences in the sample using the at least one k-mer determined as primer
Implementation Method 2
amplifying the pathogen DNA sequences in the sample using the at least one k-mer determined as primer
Data Source
Figure 1
Figure 2
Figure 3
AI summary
The present invention relates to a method of selectively amplifying at least one nucleic acid sequence of at least one microorganism and/or virus in a sample of a subject, wherein k-mers (3) are applied that show a difference in frequency and/or context in the genome (2) of the at least one microorganism and/or virus compared to the genome of the subject (1).