3D Lattice Microarray for Direct Pathogen DNA Detection
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Solution Overview
Problem
Current methods for DNA-based pathogen analysis in plant, agriculture, and food/water materials face challenges such as time-consuming and costly DNA extraction, inaccuracies in culture-based testing, interference from plant constituents, and high costs associated with chemical consumables and skilled labor, while existing DNA microarray techniques are limited by steric constraints and high energy requirements.
Innovation Solution
A 3-dimensional lattice microarray system using chemically activatable or non-covalent adsorptive solid supports with bifunctional polymer linkers and nucleic acid probes, allowing for covalent or non-covalent attachment of probes, enabling efficient DNA detection and analysis with reduced processing steps and chemical usage.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If DNA extraction is performed prior to PCR analysis, then detection accuracy is improved, but processing time and cost increase
Solution Approach 1:
The patent removes the DNA extraction step from the workflow by enabling direct PCR amplification on intact plant material. The solid support system allows PCR reagents to access and amplify target DNA sequences directly within the plant tissue matrix, eliminating the need to extract and purify DNA beforehand, thus resolving the contradiction between detection accuracy and processing time.
Solution Approach 2:
The patent segments the plant tissue into small particles or uses localized sampling spots on the solid support, allowing PCR reagents to penetrate and access DNA sequences distributed throughout the tissue. This segmentation enables direct amplification without requiring complete DNA extraction, reducing processing time while maintaining detection accuracy through distributed target access.
2Loss of time
If Colony PCR is used to eliminate extraction step, then processing time is reduced, but sensitivity decreases due to interference from specimen constituents
Solution Approach 1:
The patent introduces a specialized solid support system as an intermediary between the plant specimen and PCR reagents. This solid support provides a controlled microenvironment that facilitates DNA amplification while physically or chemically managing the interference from plant constituents, thereby maintaining high sensitivity without requiring time-consuming extraction steps.
Solution Approach 2:
The patent creates localized reaction zones on the solid support where PCR amplification occurs in confined spaces with optimized reagent concentrations. This local quality control allows the PCR reaction to proceed with high sensitivity in specific micro-environments, even when surrounded by interfering plant constituents, thus resolving the sensitivity issue while maintaining rapid processing.
3Device complexity
If direct PCR is performed on plant matter, then processing steps are reduced, but PCR amplification is heavily inhibited by plant constituents
Solution Approach 1:
The solid support acts as a mediator that separates the PCR reaction from the bulk plant matrix. It provides a controlled interface where amplification can occur with optimized conditions, protecting the reaction from inhibition by plant constituents while still allowing direct processing of intact plant material, thus maintaining both simplicity and reliability.
Solution Approach 2:
The patent modifies the physical or chemical parameters of the solid support surface to create favorable conditions for PCR amplification. By adjusting parameters such as surface chemistry, porosity, or reagent loading, the system optimizes amplification efficiency directly on plant material without requiring extraction, resolving the contradiction between process simplicity and amplification reliability.
4Measurement precision
If conventional microarray techniques are used, then DNA detection capability is achieved, but steric constraints and high energy requirements limit efficiency
Solution Approach 1:
The patent transitions from conventional two-dimensional microarray surfaces to a three-dimensional solid support structure. This dimensional change increases the effective surface area and porosity, allowing better penetration of PCR products and reducing steric constraints. The 3D architecture enables more efficient hybridization with lower energy requirements while maintaining detection capability.
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
The system provides accurate, specific, and reliable multiplex DNA detection with reduced chemical and labor costs, enabling faster results and improved sensitivity for pathogen identification in complex samples without the need for extensive sample preparation.
Implementation Method 1
a solid support having chemically activatable groups on its surface, a plurality of bifunctional polymer linkers covalently attached to the solid support
Implementation Method 2
a solid support having no chemically activatable groups on its surface, a plurality of bifunctional polymer linkers attached to the solid support by non-covalent adsorption
Implementation Method 3
a plurality of nucleic acid probes covalently attached to the bifunctional polymer linkers
Data Source
AI summary
Provided herein is a 3-dimensional lattice microarray system for DNA sequence detection and analysis. The system has a plurality of bifunctional polymer linkers, on one end of which are attached nucleic acid probes where each have a sequence complementary to signature nucleotide sequences in pathogens, plants or animals. The other end of the bifunctional polymer linker is attached to a solid support by non-covalent or covalent means. Each of the nucleic acid probes have terminal thymidine bases at the 5′ and 3′ ends that permit attachment of the probes to the bifunctional polymer linkers. Also provided is a method for fabricating the microarray system by first attaching the bifunctional polymer linkers to the solid support, followed by photochemical coupling of the nucleic probes to the microarray. A customizable microarray kit is provided that contains the solid support, linkers, probes, solvent mixture and instructions to use the kit.