Genetyping method based on SNP site nucleic acid mass spectrum detection and application thereof

Abstract

The invention relates to a genetyping method based on SNP site nucleic acid mass spectrum detection and application thereof. The genetyping method comprises: (1) designing a specific primer with respect to sequence SNP site of a target gene; (2) amplifying a target segment containing the specific SNP site of the target gene utilizing single-pipe multi-PCR; (3) digesting PCR product segment utilizing restriction enzyme; (4) performing single basic group extension reaction; (5) desalting and purifying; and (6) detecting and analyzing the gene sequence of the specific SNP site of the target gene.The genetyping method based on SNP site nucleic acid mass spectrum detection is high in detection accuracy, high in experiment repetition, large in flux, and low in cost. The invention also disclosesapplication of the genetyping method based on SNP site nucleic acid mass spectrum detection in single nucleotide polymorphism. The method can specifically and simply type the target gene or pathogenic microorganism, thereby greatly improving the typing efficiency.

Description

technical field [0001] The invention relates to the technical field of biological detection, in particular to a genotyping method based on SNP site nucleic acid mass spectrometry detection and its application. Background technique [0002] Genotyping (Genotyping) is the use of biological detection methods to determine the individual genotype (Genotype) technology. Also known as genotypic assay, the techniques used include polymerase chain reaction (PCR), DNA fragment analysis, oligonucleotide probes (ASO probes), gene sequencing, nucleic acid hybridization, gene chip technology, restriction fragments Length polymorphism (restriction fragment length polymorphism, abbreviated as RFLP) technology, etc. These techniques have been widely used in genotyping and detection of pathogenic microorganisms. But its shortcomings are also obvious, one is low throughput, and the other is that some genotypes cannot be detected, especially the detection of drug resistance of pathogenic micr...

AI Technical Summary

Problems solved by technology

Therefore, there are high requirements for the length, complexity, and reaction conditions of the sequence, and it is easily affected by various chemical factors, resulting in deviations in the detection results.

Taking the fluorescent PCR method as an example, once the gene mutation site appears in the primer hybridization region, it may cause the probe to fail to bind to the template correctly, resulting in errors in the detection results

In the gene chip, there are also mutation sites in the probe that cause false negatives in the hybridization results, and even cause incorrect pairing during the second hybridization, resulting in waste chips and affecting the detection results.

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