Host Nucleic Acid Depletion for One-Pot Pathogen Sequencing
Find Innovative SolutionsGenerate Solutions
Solution Overview
Problem
Existing methods for detecting non-host species in host samples face challenges such as high host nucleic acid background interference, loss of non-host nucleic acid signals during centrifugation, and inefficient simultaneous amplification of DNA and RNA libraries, leading to increased costs and processing time.
Innovation Solution
A method involving magnetic particles coupled to enzymes that degrade both DNA and RNA, followed by a magnetic field removal, combined with a one-pot process for creating DNA and RNA libraries, using primers to prime both single-stranded DNA and RNA, and applying lysing agents to selectively lyse biological complexes.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Object-affected harmful factors
If high-speed centrifugation is used to remove host signals, then host nucleic acid background is reduced, but non-host nucleic acid signals are lost
Solution Approach 1:
The patent applies different processing conditions to different components: mild detergent for selective lysis of host complexes, magnetic particles coupled to nucleases for targeted degradation of host nucleic acids, and controlled centrifugation parameters. This localized differentiation allows removal of host background while preserving non-host signals.
Solution Approach 2:
The patent modifies physical and chemical parameters including centrifugation speed (low speed to preserve non-host organisms), detergent concentration (mild conditions for selective lysis), and nuclease activity (controlled degradation). These parameter changes enable selective removal of host nucleic acids while maintaining non-host signal integrity.
2Quantity of substance
If separate DNA and RNA library preparation is performed, then comprehensive nucleic acid coverage is achieved, but processing time and cost increase
Solution Approach 1:
The patent combines separate DNA and RNA library preparation procedures into a single integrated workflow. Magnetic particles coupled to nucleases simultaneously process both nucleic acid types, and a unified amplification protocol handles both DNA and RNA templates, reducing processing steps while maintaining comprehensive coverage.
Solution Approach 2:
The patent employs universal reagents and protocols that work with both DNA and RNA: magnetic particles coupled to nucleases that degrade both nucleic acid types, primers that can prime both single-stranded DNA and RNA, and a single amplification protocol. This multi-functionality eliminates the need for separate processing lines.
3Quantity of substance
If conventional library preparation methods are used with low nucleic acid amounts, then sample integrity is maintained, but processing complexity and cost increase
Solution Approach 1:
The patent employs self-service mechanisms where magnetic particles automatically capture and concentrate nucleic acids from complex matrices, and nucleases selectively degrade host background without requiring complex purification steps. The system uses its own components (magnetic particles, nucleases, primers) to perform multiple functions sequentially.
Solution Approach 2:
The patent extracts the essential function of library preparation by using magnetic particles to selectively bind and concentrate target nucleic acids, then directly proceeding to amplification without complex intermediate purification steps. This extraction of key functions simplifies the overall procedure while maintaining sample integrity.
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 effectively degrades unwanted nucleic acids, enhances signal-to-background ratio, and enables rapid, cost-effective creation of libraries for accurate detection of non-host species, facilitating widespread use of next-generation metagenomic sequencing.
Implementation Method 1
applying a magnetic field to remove the magnetic particles coupled to enzymes from the sample
Implementation Method 2
magnetic particles coupled to enzymes such as nucleases to degrade unwanted nucleic acids present in the sample
Implementation Method 3
applying lysing agents to selectively lyse biological complexes
Data Source
AI summary
The present invention provides a method of detecting non-host species in a host sample. The host sample is contacted with magnetic particles coupled to enzymes to degrade cell-free host nucleic acids present in the host sample, wherein the magnetic particles coupled to enzymes are capable of degrading both DNA and RNA. A magnetic field is applied to remove the magnetic particles coupled to enzymes from the host sample. A DNA and RNA library is created from the degraded sample in a one-pot process, followed by detection of the presence of non-host nucleic acids from the DNA and RNA library.