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Kit for detecting mutation of mitochondria T12201C related with epicophosis and application

A T12201C, mitochondrial technology, applied in the determination/inspection of microorganisms, biochemical equipment and methods, etc., can solve problems such as trouble, achieve the effect of alleviating pain, intuitive interpretation of results, and simple detection process

Active Publication Date: 2014-12-17
ZHEJIANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This blood collection method has brought some pain and injury to many subjects, especially infants and young children, and there are certain risks and troubles when transferring samples across regions

Method used

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  • Kit for detecting mutation of mitochondria T12201C related with epicophosis and application
  • Kit for detecting mutation of mitochondria T12201C related with epicophosis and application
  • Kit for detecting mutation of mitochondria T12201C related with epicophosis and application

Examples

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Effect test

Embodiment 1

[0037] Embodiment 1 A kit of the present invention

[0038] see figure 1 , the kit provided by the present invention consists of DNA extraction mixture 1, PCR mixture 2 for amplifying T12201C fragment, a pair of outer primers 3 designed for T12201C, inner primer 4 designed for T12201C, restriction endonuclease 5, positive Control 6, negative control 7 and box body 8 are composed, wherein, DNA extraction mixed solution 1 is mainly made up of cell lysate, proteinase K solution, chloroform, phenol, absolute alcohol; PCR mixed solution 2 that amplifies T12201C fragment comprises dNTP ( deoxymononucleotide), 10×PCR buffer, MgCl 2 , triple distilled water and Taq Enzyme (DNA polymerase), outer primer 3 designed for T12201C has outer forward primer F: TAT CAC TCT CCT ACT TAC AG (SEQ ID NO: 1), outer reverse primer R: AGA AGG ATA TAA TTC CTA CG ( SEQ ID NO:2); Inner primer 4 designed for T12201C includes inner forward primer F: AATATAGTTTAACCAAAACATCAGATTGTGAATCTGACAACAGAGGCTTACGA...

Embodiment 2

[0040] Example 2 Carrying mitochondrial tRNA Ala Detection of deaf families with T12201C mutation

[0041] 1. Test samples

[0042] A deaf family carrying the T12201C mutation was selected. See his pedigree Figure 4 . This family presents a typical maternal inheritance, and all patients have deafness as the only clinical symptom, but the blood pressure of each affected member in the family varies. The family has a total of 88 members, including 35 members of the maternal line and 17 deaf persons.

[0043] 2. Extraction of Genomic DNA

[0044] Samples from 5 subjects were collected respectively (including the capillary blood drop of Ⅲ-6 and Ⅲ-9; Ⅲ-13 was oral mucosa scraping or saliva; Ⅳ-29 and Ⅳ-31 were blood follicles hair), cut the blood filter paper into pieces of about 1cm2 with clean scissors, put them into a 1.5ml EP tube (Eppendorf tube), add 900μl red blood cell lysis buffer (20mmol / L Tris-HCl, pH 7.6) and let it stand at room temperature Set it for 10 minutes...

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Abstract

The invention aims to provide a kit for detecting mutation of mitochondria T12201C related with epicophosis, and the kit consists of a DNA extraction mixed liquor, a PCR mixed liquor for amplifying T12201C segments, a pair of outer primers designed according to T12201C, a pair of inner primers designed according to T12201C, restriction enzyme, a positive control unit, a negative control unit and a kit body. By using a proteinase K digestion cracking method to extract the genomic DNA from a few samples, the kit disclosed by the invention not only solves the problem that the DNA is extracted from blood of an epicophosis patient traditionally, relieves the pain of the detected patient, but also solves the problem of sample transmission between different areas, not only improves the specificity of enzyme digestion but also ensures the specificity and stability of the results, not only lowers the detection cost but also realizes simplicity and quickness in operation, and can be applied to detection of mutation of mitochondria gene T12201C related with epicophosis.

Description

technical field [0001] The invention belongs to the field of biological technology, and relates to a kit for detecting the mitochondrial DNA T12201C mutation related to deafness, and also relates to a detection method for the mitochondrial gene mutation related to deafness, in particular to detecting mitochondrial tRNA His The method for the T12201C mutation, and the application of the above-mentioned method or the above-mentioned kit in detecting the mitochondrial DNA T12201C mutation related to deafness. Background technique [0002] Deafness is a common disabling disease that causes speech communication barriers and is one of the greatest sufferings of human beings. According to WHO estimates in 2005 that there were 278 million hearing disabled people in the world, accounting for 4.6% of the world's total population; 80% of hearing disabled people live in low- and middle-income countries. Deafness has become a serious health and safety problem in my country, bringing a h...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68
Inventor 管敏鑫蒋萍萍冀延春郑静梁敏龚莎莎徐静
Owner ZHEJIANG UNIV
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