PCR Primer Design for Mycobacterium Detection Sensitivity

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Solution Overview

Problem

Current methods for detecting Mycobacterium tuberculosis complex (MTBC) in early stages of infection are inadequate, leading to high mortality rates if treatment is not initiated promptly, highlighting the need for a reliable and efficient detection method.

Innovation Solution

A method and kit utilizing specific primers and probes, such as SEQ ID NOs. 1, 2, and 3, for performing PCR to amplify and detect the IS6110 sequence in clinical samples like blood, sputum, or bronchoalveolar lavage fluid, enabling early detection of MTBC through real-time quantitative PCR.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Measurement precision

If conventional detection methods are used for Mycobacterium tuberculosis complex, then the detection process is simple, but the detection sensitivity and precision are insufficient leading to high mortality rates

Engineering Contradiction:

Improvedetection sensitivityVSAvoiddetection method complexity

Core Design Contradiction:

Measurement precisionVSDevice complexity

Solution Approach 1:

The patent segments the detection process into multiple specific steps: DNA extraction from clinical samples, PCR amplification using specifically designed primers (SEQ ID NOs: 1 and 2) targeting the IS6110 sequence, and electrophoresis analysis. This segmentation allows each step to be optimized independently, achieving high detection sensitivity (detecting as few as 5 bacterial copies) while maintaining procedural clarity and manageability

Inventive Principle:

Principle #1Segmentation

Solution Approach 2:

The patent performs preliminary action by designing and pre-validating specific primer sequences (SEQ ID NOs: 1 and 2) that target the IS6110 sequence unique to Mycobacterium tuberculosis complex. These primers are optimized beforehand to ensure high specificity and sensitivity, allowing rapid detection without requiring complex real-time monitoring equipment. The primers are ready for immediate use in PCR amplification, eliminating the need for complex instrument setup during actual detection

Inventive Principle:

Principle #10Preliminary action

2Reliability

If treatment is delayed until later stages of tuberculosis infection, then the treatment complexity is reduced, but the mortality rate increases to about half within three years

Engineering Contradiction:

Improvemortality reductionVSAvoiddetection time

Core Design Contradiction:

ReliabilityVSLoss of time

Solution Approach 1:

The patent replaces complex mechanical/digital real-time PCR systems with a simpler biochemical amplification system using conventional PCR and electrophoresis. The specifically designed primers (SEQ ID NOs: 1 and 2) enable high-sensitivity detection through biochemical amplification of the IS6110 sequence, which can be visualized using standard electrophoresis equipment rather than requiring expensive real-time thermal cyclers with fluorescence detection capabilities. This substitution reduces equipment complexity while maintaining high detection reliability

Inventive Principle:

Principle #28Mechanics substitution (Replace mechanical system)

Solution Approach 2:

The patent changes the detection parameter from requiring complex real-time fluorescence monitoring to using conventional end-point PCR with electrophoresis visualization. By optimizing the primer sequences (SEQ ID NOs: 1 and 2) and PCR conditions, the method achieves high sensitivity (detecting 5 copies) using standard laboratory equipment, thereby reducing both time and cost while improving reliability for early tuberculosis detection

Inventive Principle:

Principle #35Parameter changes

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 method achieves sensitive and specific detection of MTBC, with a minimum detection limit of 5 copies, improving the chances of timely treatment and reducing mortality by enabling early identification of infected patients.

Implementation Method 1

performing a polymerase chain reaction (PCR) by using the primers and the sample to obtain a product

Methodology Applied

Scientific EffectPolymerase chain reaction (PCR):

Implementation Method 2

providing a pair of primers, wherein the primers are selected from the group consisting of SEQ ID NO. 1, SEQ ID NO. 2

Methodology Applied

Scientific EffectDNA hybridization:

Data Source

PatentUS11174520B2Method for detecting presence or absence of <i>Mycobacterium </i>and kit thereof

Publication Date: 2021.11.16 DELTA ELECTRONICS INC(CN)

AI summary

A method and a kit for detecting mycobacterium are provided. The method includes steps of: providing a sample; providing a pair of primers, which is selected from a group consisting of a sequence having about 45% to 100% similar to SEQ ID NO. 1, a sequence having about 60% to 100% similar to SEQ ID NO. 2, a sequence complementary thereof; performing a polymerase chain reaction by using the set of primers and the sample to obtain a product; and analyzing the product to detect the presence of the mycobacterium.