HLA-B*15:02 Detection via Single SNP Locus Fluorescent PCR
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Solution Overview
Problem
Current methods for detecting the HLA-B*15:02 allele, such as multiplex real-time fluorescent PCR, are complex, have limited specificity, and cannot distinguish between homozygotes and heterozygotes, making them inefficient and inaccurate.
Innovation Solution
A specific primer probe combination targeting a single SNP locus, rs144012689, is used in a fluorescent PCR method, combined with an internal reference gene, to accurately detect HLA-B*15:02 alleles, enabling differentiation between homozygotes and heterozygotes with high specificity and efficiency.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If multiplex real-time fluorescent PCR method is used to detect HLA-B*15:02 alleles, then detection throughput is improved, but method complexity increases and specificity is limited
Solution Approach 1:
The patent extracts and focuses on a single specific SNP locus (rs144012689) from the complex HLA-B gene region, designing primers and probes that target only this critical locus. This extraction approach simplifies the detection method while maintaining high throughput capability, resolving the contradiction between complexity and productivity
Solution Approach 2:
The patent segments the HLA-B gene detection into a specific functional unit by targeting only the rs144012689 SNP locus with dedicated primers and probes. This segmentation allows for streamlined reaction conditions and simplified analysis while preserving the ability to perform high-throughput screening
2Adaptability or versatility
If multiple SNP loci are used to identify HLA-B*15:02, then detection coverage is improved, but final specificity is limited and homozygote/heterozygote differentiation is impossible
Solution Approach 1:
The patent applies local quality by concentrating all detection specificity into one critical SNP locus (rs144012689) rather than distributing it across multiple loci. The primers and probes are designed with specific sequence characteristics that enable both high coverage and precise differentiation of homozygous and heterozygous states at this single locus
Solution Approach 2:
The patent uses fluorescent probes with distinct emission wavelengths (different 'colors') that bind specifically to different genotypes at the rs144012689 locus. This allows simultaneous detection and differentiation of various genotype states through fluorescence signal analysis, achieving both high coverage and precision
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 high throughput, high specificity, low cost, and rapid detection of HLA-B*15:02 alleles, with the ability to differentiate between homozygotes and heterozygotes, improving detection efficiency and accuracy compared to existing methods.
Implementation Method 1
Fluorescent Quantitative PCR (Real time-PCR)
Data Source
AI summary
Disclosed is a fluorescent PCR method for detecting HLA-B*15:02 allele and a specific primer probe combination. In the present disclosure, a set of primers and probes are designed based on an HLA-B*15:02 specific SNP gene locus by using TaqMan probe technology, combining another set of primers and probes corresponding to the internal reference gene β-Actin, and a set of primer probe for non-HLA-B*15:02 genes are designed to detect whether a DNA sample contains an HLA-B*15:02 gene and whether a sample is homozygous or heterozygous. Compared with the similar detection methods in the past, the technical scheme in the present disclosure inherits the advantages of high specificity, high throughput, high resolution, low cost, simple and convenient operation, process controllability and the like of the fluorescent PCR, and may detect whether a sample is homozygous or heterozygous.