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Mitochondria DNA11778 point mutation detecting reagent case by molecular probe technology

A molecular probe and technical detection technology, which is applied in biological testing, material inspection products, microbial measurement/inspection, etc., can solve the problem of low SNP recognition rate, and achieve the effect of simple operation, short time-consuming and good accuracy

Active Publication Date: 2009-02-04
ZHEJIANG UNIV +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Therefore, when using TaqMan MGB fluorescent probes to detect SNPs, the length of the probes is generally in the range of 13 to 18 bases, while the length of conventional TaqMan probes will reach 30-40 bases when the content of AT bases is large. The longer the needle length, the lower its SNP recognition rate

Method used

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  • Mitochondria DNA11778 point mutation detecting reagent case by molecular probe technology
  • Mitochondria DNA11778 point mutation detecting reagent case by molecular probe technology
  • Mitochondria DNA11778 point mutation detecting reagent case by molecular probe technology

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Experimental program
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specific Embodiment 1

[0038] The raw materials for preparing the kit of the present invention are selected as follows:.

[0039] PCR primers were synthesized by Biosearch.INC, and the primer sequences were:

[0040] MT-1: 5'-CATCCTCATTACTATTCTGCCTAGCA-3';

[0041] MT-2: 5'-GGAGTAGAGTTTGAAGTCCTTGAGAGA-3';

[0042]Molecular probe: design a molecular beacon (MGB) probe complementary to the target sequence in the conserved region of the cell genome, label the 5' end with FAM, and the other end with MGBNFQ ((Minor groove binder / Non-fluorescent quencher)), which The sequence is:

[0043] MT-3: 5'-FAM-CACTCACAGTCACATCA-MGBNFQ;

[0044] MT-4: 5'-VIC-AGGATTATGATGCGACTGT-MGBNFQ.

[0045] dNTPs (dATP, dTTP, dnP, dGTP): purchased from APPlied Biosystems, concentration: 200 μMol / L.

[0046] The kit components in the specific embodiment 1 are as follows:

[0047] 2×Taqman PCR buffer half of the total volume of the reaction;

[0048] Each 200μMol / L dATP, dTTP, dGTP, dCTP;

[0049] Two primers MT-1 and MT-...

specific Embodiment 2

[0053] The kit components are as follows:

[0054] 2×Taqman PCR buffer half of the total volume of the reaction;

[0055] Each 200μMol / L dATP, dTTP, dGTP, dCTP;

[0056] Two primers MT-1 and MT-2 of 2 μMol / L each;

[0057] Two probes MT-3 and MT-4 of 5 μMol / L each;

[0058] 2.5mMol / L magnesium chloride;

[0059] The balance is distilled water.

specific Embodiment 3

[0060] The kit components are as follows:

[0061] 2×Taqman PCR buffer half of the total volume of the reaction;

[0062] Each 200μMol / L dATP, dTTP, dGTP, dCTP;

[0063] Two primers MT-1 and MT-2 of 1 μMol / L each;

[0064] Two probes MT-3 and MT-4 at 2.5 μMol / L each;

[0065] 2mMol / L of magnesium chloride;

[0066] The balance is distilled water.

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Abstract

The invention relates to a gene testing method and its agent box, which provides a based molecular fluorescence, probe PCR mitochondria DNA11778 point mutation measuring method and its agent box. The invention first quotes weather existing mitochondria DNA11778 point mutation, and then designs lead substance at mitochondria genome conservation zone and molecular probe which is complemented with target series at mitochondria 11778 point, and then dose PCR growth process.

Description

technical field [0001] The invention relates to a kit for gene detection, more specifically, the invention relates to a kit for detecting mitochondrial DNA11778 point mutation by PCR based on molecular fluorescent probe technology. Background technique [0002] Leber's disease, also known as Leber's hereditary optic neuropathy (LHON), is a hereditary disease that mainly affects the retina and the fibers of the papillary macular tract at the front of the scleral cribriform plate, leading to degeneration of the optic nerve. The mode of transmission of the disease does not conform to Mendelian inheritance, and the mode of inheritance and pathogenesis are still controversial. Clinically, it mostly occurs in young and middle-aged adults, and both eyes present acute or subacute severe damage to visual function at the same time or successively. and patients with many adverse consequences. Clinically carrying out LHON definite molecular genetic diagnosis can not only avoid expensi...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N21/76G01N33/48C12Q1/68
Inventor 王剑勇李毓敏王竞薛忆邵俊斌
Owner ZHEJIANG UNIV