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High-throughput single nucleotide polymorphism detecting method based on magnetic nano-particles and universal label technique

A magnetic nanoparticle, high-throughput technology, applied in the field of genetic testing, can solve the problems of high price and increase the cost of a large number of loci typing, and achieve the effect of reducing costs

Inactive Publication Date: 2008-06-25
李松 +1
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The method of SNPs typing by fluorescent probe hybridization method needs to design two labeled probes for each SNPs site, and the labeled probes are expensive, which greatly increases the cost of typing a large number of sites

Method used

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  • High-throughput single nucleotide polymorphism detecting method based on magnetic nano-particles and universal label technique

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Embodiment approach I

[0030]The invention relates to a method for high-throughput SNPs typing by using magnetic nanoparticles as carriers and universal label technology. The details are as follows: 1. Select several important functional SNP sites, design corresponding primers (one side of the primer is biotin-labeled), a pair of wild and mutant detection probes 4, 6 (sequences include those complementary to the target sequence, etc.) Gene-specific sequence, a Poly T spacer sequence with 11 bases in the middle and a universal label sequence) and a pair of labeled (fluorescent, nano-gold, luminescent enzyme) universal label detection probes 3, 5 (general The length of the detection probe is about 13 bases, which is complementary to the universal label sequence of the detection probe). 2. Amplify the biotin-labeled target sequence containing the SNPs site to be tested by PCR. The biotin-labeled target sequence can be the product of conventional PCR amplification, or it can be solid-phase amplification...

Embodiment approach II

[0032] The details are as follows: 1. Select several important functional SNP sites, design corresponding primers (one side of the primer is marked), a pair of wild and mutant detection probes 4, 6 (sequences include alleles complementary to the target sequence) specific sequence, a Poly T spacer sequence of 11 bases in the middle and a universal label sequence) and a pair of two-color fluorescent-labeled (wild-type Cy3, mutant Cy5) universal label detection probes 3 and 5 (universal detection probes The length is about 13 bases, and it is complementary to the universal label sequence of the detection probe). 2. Amplify the marked target sequence containing the SNPs site to be tested by PCR. The marked target sequence can be the product of conventional PCR amplification, or the product of solid-phase amplification PCR on the surface of the magnetic nanoparticles. 3. If the labeled target sequence 2 is a conventional amplification product, it is necessary to add an appropriate ...

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Abstract

The invention discloses a multi-sample multi-site typing method against single nucleotide polymorphisms (SNP) in a genome by using magnetic nano particles as carriers and combining with the technology of universal label probe. The invention is characterized in that: a labeled PCR product is directly fixed on the magnetic nano particles through covalent unions between labels, or a labeled primer is fixed on the magnetic nano particles to directly execute solid-phase PCR amplification, thereby forming magnetic nano particles-DNA complex (MNP-ssDNA). Then, the MNPs-ssDNA is hybridized with a detecting probe of corresponding site and a labeled universal label probe, thereby achieving multi-sample multi-site high flux SNP detection. The advantages of the invention are that: only a pair of labeled probes are required by using the universal label technology to achieve the multi-site typing, and the typing cost is greatly reduced; meanwhile, the invention overcomes the defects of high cost, time consuming, taking trouble, etc. caused by purifying and concentrating a target sequence waiting for detecting in the prior art by using the advantages that the magnetic nano particles are easily separated, etc. Therefore, the method has the advantages of typing mass samples simply, efficiently and accurately.

Description

Technical field [0001] The invention relates to a gene detection technology using magnetic nanoparticles as a carrier combined with a universal label probe, especially a single nucleotide suitable for automation, multi-site, multi-sample, high-throughput, and low-cost Polymorphism (Single Nucleotide Polymorphisms, SNPs) detection method. Background technique [0002] With the completion of the Human Genome Project (HGP), gene sequencing provides a platform for studying the secrets of genetic information, including the study of human genomes, which greatly facilitates the study of gene families. research etc. The establishment of SNPs markers not only facilitates the cloning of the causative genes of genetic diseases, but also facilitates the research of various genetic diseases such as single-gene and polygenic diseases. SNPs markers are also of great significance to clinical medicine. SNPs markers serve clinical medicine as an effective genetic polymorphism marker, and ca...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68
Inventor 李松何农跃刘洪娜田岚
Owner 李松
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