Nucleic Acid Amplification via Nonionic Surfactant and Temperature Control
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Solution Overview
Problem
Existing nucleic acid amplification and detection methods are inefficient and time-consuming, particularly in detecting multiple types of viruses from various specimens.
Innovation Solution
A nucleic acid amplification method and system that utilizes a nonionic surfactant, dNTPs, and NTPs, along with temperature-controlled incubation steps and primers, to amplify nucleic acids, followed by fluorescence detection using probe DNA with immobilized fluorescent and quenching molecules.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If conventional nucleic acid amplification methods are used, then detection sensitivity can be achieved, but the detection process is time-consuming and inefficient
Solution Approach 1:
The patent combines multiple separate steps (nucleic acid extraction, amplification preparation, and amplification reaction) into a single integrated reaction tube. The extraction buffer contains all necessary components (surfactants, proteins, enzymes) to perform extraction and amplification simultaneously without transferring samples between tubes, thereby reducing time while maintaining sensitivity
Solution Approach 2:
The extraction buffer is pre-formulated with optimized concentrations of nonionic surfactants (Tween 20, Triton X-100), ionic surfactants (SDS), proteins (BSA, lysozyme), and enzymes (RNase A, proteinase K) before the reaction begins. This preliminary preparation eliminates the need for separate extraction steps and enables immediate amplification, significantly reducing detection time
2Adaptability or versatility
If multiple types of viruses are detected from various specimens, then comprehensive detection capability is achieved, but the process becomes inefficient and time-consuming
Solution Approach 1:
The extraction buffer is designed as a universal reagent that can process various specimen types (throat swabs, nasal swabs, sputum, saliva, urine, stool) and detect multiple virus types simultaneously. The buffer contains broad-spectrum components that work across different sample matrices and virus nucleic acids, enabling one-step processing for diverse detection needs
Solution Approach 2:
The patent segments the detection process into distinct functional components within the single tube: extraction components (surfactants, proteins, enzymes) separated from amplification components (primers, dNTPs, buffer). This segmentation allows each component to be optimized independently while working together efficiently, maintaining versatility across different virus types without compromising productivity
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
Enables rapid and sensitive detection of multiple viruses in various specimens with high sensitivity, reducing the time required for nucleic acid extraction and amplification.
Implementation Method 1
adding a nonionic surfactant, dNTPs, and NTPs to a specimen containing a virus
Implementation Method 2
retaining the first liquid in a first temperature zone for a first period of time
Implementation Method 3
adding a first primer corresponding to a first region of a nucleic acid molecule derived from the virus, a second primer corresponding to a second region different from the first region, and an enzyme to the first liquid
Implementation Method 4
retaining the second liquid in a second temperature zone for a second period of time
Implementation Method 5
Each molecule of the probe DNA is modified with a fluorescent molecule and a quenching molecule. A distance between the fluorescent molecule and the quenching molecule in the probe DNA not bound to the nucleic acid molecule amplified by the enzyme is shorter than a distance between the fluorescent molecule and the quenching molecule in the probe DNA bound to the nucleic acid molecule amplified by the enzyme
Data Source
AI summary
To achieve a nucleic acid amplification method and a nucleic acid amplification system capable of simply amplifying a nucleic acid in a short period of time, and a virus detection method and a virus detection system. The nucleic acid amplification method includes: a first preparation step of preparing a first liquid by adding a nonionic surfactant, dNTPs, and NTPs to a specimen containing a virus; a first incubation step of retaining the first liquid in a first temperature zone for a first period of time; a second preparation step of preparing a second liquid by adding a first primer corresponding to a first region of a nucleic acid molecule derived from the virus, a second primer corresponding to a second region, and an enzyme to the first liquid after the first incubation step; and a second incubation step of retaining the second liquid in a second temperature zone for a second period of time.