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Mononucleotide polymorphism (SNP) high-pass typing method based on magnetic nano particle polymerase chain reaction (PCR)

A magnetic nanoparticle and single nucleotide polymorphism technology, applied in the field of single nucleotide polymorphism typing, can solve the problems of high cost, inability to give sequence information, high operator requirements, etc., and achieve positive mismatch The effect of high number ratio, intuitive typing results and high application value

Inactive Publication Date: 2008-03-12
刘洪娜 +1
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Problems solved by technology

For example, detection methods based on different molecular conformations can only detect mutations, but cannot give sequence information, and cannot determine the specific mutation site and mutation form. At the same time, such methods have high requirements for operators.
However, various detection methods based on fluorescence resonance energy transfer still have disadvantages such as high cost or low throughput.
Most of the SNP typing methods based on molecular hybridization technology need to purify and concentrate the PCR products containing SNP sites, which is a time-consuming and labor-intensive task

Method used

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  • Mononucleotide polymorphism (SNP) high-pass typing method based on magnetic nano particle polymerase chain reaction (PCR)

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Embodiment Construction

[0022] The invention relates to a high-throughput SNPs detection method. The details are as follows: 1. Polymerase chain reaction based on magnetic nanoparticles: biotin-labeled primers are immobilized on avidin-coated magnetic nanoparticles, and carried out in a system containing free primers, dNTPs, polymerase and template DNA The PCR reaction directly amplifies the target fragment (MNPs-DNA) containing the detected SNP site on the surface of the magnetic nanoparticle. 2. Denaturation and quenching, the double-stranded PCR product on the surface of the magnetic particle becomes single-stranded (MNPs-ssDNA). 3. Hybridization: Design a pair of allele-specific detection probes (wild-type Cy3 marker, mutant Cy5 marker) according to the sequence at the gene site to be tested, and the ssDNA immobilized on the surface of the particle and the designed probe are Hybridization at a specific temperature. 4. Fully wash to remove the free target sequence, denature, spot the denatured f...

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Abstract

The invention discloses a new method for parting the single nucleotide polymorphisms (SNP) taking use of magnetic nanometer particle polymerase chain reaction (PCR), which is characterized in that a primer at a single side is fixed on magnetic nanometer particles and target sequence is expanded directly on the surfaces of magnetic nanometer particles, and then the sample is parted by hybridizing magnetic nanometer particles-ssDNA complex with allele specific probes. The method needs no purify and concentrate the product, the scans and parts rapidly and straightforwardly, is a parting method that is easy and rapid to operate, and is of high throughput and sensibility.

Description

Technical field [0001] The invention relates to a novel typing method of single nucleotide polymorphism. The method of combining magnetic nanoparticle polymerase chain reaction (MNPs-PCR) and allele-specific hybridization is used to realize the typing of functional SNP loci in a large number of samples. On the basis of high throughput, convenience, applicability and high reliability, the technology greatly simplifies the operation steps. Background technique [0002] Single nucleotide polymorphism (SNP) refers to the presence of two different bases at a specific nucleotide position in the genome, at least one of which has a frequency of not less than 1% in the population. Although the genetic code consists of 4 bases, a SNP is usually just 1 biallelic or dimorphic genetic variation. SNPs are the most common variations in human gene sequences, and their study helps to explain individual phenotypic differences, susceptibility of different groups and individuals to complex di...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68
Inventor 刘洪娜何农跃李松贺全国陆祖宏林琳
Owner 刘洪娜
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