Kit for detecting mutation of duchenne muscular dystrophy causing gene and use thereof

A technology for muscular dystrophy and disease-causing genes, which is applied in the field of kits for detecting mutations in the disease-causing genes of pseudohypertrophic muscular dystrophy, and can solve problems such as unclear pathogenesis of pseudohypertrophic muscular dystrophy , to reduce the incidence of

Inactive Publication Date: 2011-11-02
上海佰真生物科技有限公司 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The pathogenesis of pseudohypertrophic muscular dystrophy is unclear, and there is no effective treatment so far

Method used

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  • Kit for detecting mutation of duchenne muscular dystrophy causing gene and use thereof
  • Kit for detecting mutation of duchenne muscular dystrophy causing gene and use thereof
  • Kit for detecting mutation of duchenne muscular dystrophy causing gene and use thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] A kit for detecting whether there is a mutation in the pathogenic gene DMD of pseudohypertrophic muscular dystrophy for one person, including the following components:

[0029] (1) 79 pairs of primers for PCR amplification and sequencing of exons 1 to 79 of the DMD gene, each primer 1 OD, and the sequences of each primer are shown in Table 1;

[0030] (2) PCR amplification reagent: dNTP mixture (2.5mM each) 72μl, 10X PCR reaction buffer (Mg 2+ free)90μl, MgCl 2 (25mM) 90μl, T aq DNA polymerase (5Units / μl) 7.2μl;

[0031] (3) PCR product purification reagents: SAP enzyme (1units / μl) 33.75μl, ExoI enzyme (10units / μl) 16.875μl;

[0032] (4) Sequencing reagents, BigDye mix (25%) 90μl, EDTA solution (125mM) 90μl, ethanol solution (100%) 1.35ml, ethanol solution (70%) 2.7ml, HIDI solution 720μl.

[0033] This kit is stored at -20°C, and repeated freezing and thawing should be avoided as much as possible.

Embodiment 2

[0035] The steps for using the kit for detecting whether a mutation occurs in the pseudohypertrophic muscular dystrophy gene DMD for one person include:

[0036] (1) extract the genomic DNA of the sample;

[0037] (2) DMD gene PCR amplification:

[0038] A total of 79 PCR reactions were performed with 79 pairs of primers. Each reaction system has a total volume of 10 μl, including 1.0 μl of genomic DNA (100ng / μl), 0.8 μl of dNTP mixture (2.5mM each), 10X PCR reaction buffer (Mg 2+ free) 1.0 μl, each 0.2 μl of upstream primer and downstream primer (10 μM), MgCl 2 (25mM) 1.0μl, T aq DNA polymerase (5Units / μl) 0.08μl, deionized water 5.72μl. The reaction conditions were 94°C for 12 minutes, 30 cycles of 94°C for 30 seconds, 60°C for 30 seconds, 72°C for 30 seconds, and then 72°C for 10 minutes.

[0039] (3) PCR product purification:

[0040] A total of 79 PCR purification reactions were performed. Each reaction system has a total volume of 12.5 μl, including 10 μl of PCR pr...

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PUM

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Abstract

The invention discloses a kit for detecting if a duchenne muscular dystrophy (DMD) causing gene has a mutation. The kit comprises the following main components: (1) 79 pairs of primers for PCR amplification and sequencing of 1 to 79 extrons of the DMD causing gene; (2) a polymerase chain reaction (PCR) amplification reagent; (3) a PCR product purification reagent; and (4) a DNA sequencing reagent. The kit disclosed by the invention is used for detecting the mutation of a duchenne muscular dystrophy causing gene, can quickly and conveniently detect people or patient with the DMD causing gene, can be used for prenatal diagnosis, prevention of birth of affected children, and can lower the incidence rate of the DMD.

Description

Technical field [0001] The invention belongs to the field of biological technology, and in particular relates to a kit for detecting whether a mutation occurs in a pathogenic gene of pseudohypertrophic muscular dystrophy. Background technique [0002] Pseudohypertrophic muscular dystrophy is the most common X-linked recessive genetic disease of the nervous system. Patients present with progressive weakness and atrophy of skeletal muscles, and pseudohypertrophy of calf gastrocnemius muscles. There are two types. Duchenne muscular dystrophy (DMD) is a more serious type, with an incidence rate of 1 / 3500 live-born male babies. It usually dies of respiratory failure and / or heart failure around the age of 20. Becker's muscular dystrophy (Becker's muscular dystrophy, BMD) is a type with milder symptoms than DMD, with an incidence rate of 1 / 20000, and it does not affect the patient's life expectancy. [0003] Duehenne muscular dystrophy (Duehenne muscular dystrophy, DMD) is caused ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68
Inventor 秦胜营贺林陈培华马端陈伟坚王桢媚
Owner 上海佰真生物科技有限公司
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