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Methods and kits for detecting SNPs using universal Taqman probes

A kit and a general-purpose technology, applied in the field of SNP detection, can solve the problems of sequence specificity, low detection sensitivity, and slow signal speed

Active Publication Date: 2020-12-25
CHINA GOLDEN MARKER BEIJING BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Moreover, these detection methods and systems all have the defects of low sequence specificity, low detection sensitivity, and slow speed of signal emergence.
Although the KASP method is currently a widely used method using universal probes, the universal fluorescent probe forms a double strand with the universal quencher probe and can only be used for SNP typing detection

Method used

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  • Methods and kits for detecting SNPs using universal Taqman probes
  • Methods and kits for detecting SNPs using universal Taqman probes
  • Methods and kits for detecting SNPs using universal Taqman probes

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Experimental program
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Embodiment

[0110] Experimental materials and methods:

[0111] (1) PCR reaction primers and probes (see Table 1 below):

[0112] Table 1

[0113]

[0114]

[0115] Note 1: The underlined part of the primer is the Tail sequence.

[0116] The SNPs to be detected: the positions and flanking sequences of the A004914 and A004920 sites in the reference genome (the reference genome is version B73V2), see Table 2 below:

[0117] Table 2

[0118]

[0119] (2) Reagents required for the reaction:

[0120] Tris-HCl, KCl, Triton X-100, rox, DMSO and glycerol were purchased from Sigma; dNTPs, MgCl 2 and Taq enzymes were purchased from Vazyme biotech co.,ltd.; general TaqMan probes and primers were synthesized by Thermo Fisher Scientific Inc.

[0121] (3) PCR reaction system:

[0122] Mix1 reaction system, each PCR reaction system is 3 μl:

[0123] Reagent working concentration Tris‐HCl (pH8.4) 20mM KCl 50mM dNTPs 0.2mM MgCl 2

1.5mM Taq e...

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Abstract

The invention relates to a method for detecting SNP (single nucleotide polymorphism) by virtue of a universal TaqMan probe and a kit, in particular to a kit comprising a universal TaqMan probe formed by a universal oligonucleotide sequence with a report fluorophore and a quenching group at terminals 5' and 3' respectively and a primer formed by a universal tail sequence part and an allele specific sequence part, as well as a method for detecting the SNP by virtue of the kit.

Description

technical field [0001] The invention relates to a method and a kit for detecting SNP by using a universal TaqMan probe. In particular, it relates to a universal TaqMan probe comprising a universal oligonucleotide sequence consisting of a reporter fluorophore and a quencher group at the 5' end and a quencher group at the 3' end, respectively, and a universal tail sequence (Tail) part and an allele A kit of primers composed of specific sequence parts, and a method for detecting SNP using the kit. Background technique [0002] The methods currently commonly used to detect PCR products, especially single nucleotide polymorphisms (SNPs), mainly involve: [0003] (1) Probe method (U.S. Patent No. 5,538,848) (CAST-PCR "Competitiveallele-specific TaqMan PCR"), which detects specific PCR products by hybridizing and cleavage of a double-labeled fluorogenic probe in an amplification reaction. One or two primers need to be designed that anneal at the site of the sequence change to b...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6858C12N15/11
CPCC12Q1/6827C12Q2561/101C12Q2563/107C12Q2561/113
Inventor 翟晨光赵丽娜卢洪
Owner CHINA GOLDEN MARKER BEIJING BIOTECH CO LTD