Therapeutic effects of Math1 gene pairs forsensorineural hearing loss

A gene and adenovirus technology, applied in the field of construction of recombinant adenovirus vector for inner ear specific expression gene Math1, can solve the problems of restricting the application of stem cells and difficult separation

Inactive Publication Date: 2009-03-25
杨仕明 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although a method of isolating inner ear stem cells has been found, the difficulty of isolating stem cells limits the application of stem cells in deafness treatment. So far, gene therapy is still the most promising way to treat deafness.

Method used

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  • Therapeutic effects of Math1 gene pairs forsensorineural hearing loss
  • Therapeutic effects of Math1 gene pairs forsensorineural hearing loss
  • Therapeutic effects of Math1 gene pairs forsensorineural hearing loss

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] Embodiment 1, plasmid digestion reaction:

[0047] The appropriate reaction temperature and reaction buffer were selected according to the conditions required for various restriction endonuclease reactions. The reaction system was 20ul, and the dosage of each enzyme was 5U.

[0048]

[0049]

Embodiment 2

[0050] Embodiment 2, glass milk method reclaims DNA fragment:

[0051] After the DNA sample undergoes electrophoresis, cut off the DNA fragment to be recovered under ultraviolet light, add 3 times the volume of sol solution, bathe in 60°C water for 5 minutes, add 10ul glass milk, leave at room temperature for 5 minutes, centrifuge at 8000rpm for 1 minute, and remove the supernatant , add 125ul rinse solution, centrifuge at 8000rpm for 1 minute, remove the supernatant, then add 20ulTE buffer, mix well, bathe in 60°C water for 5 minutes, centrifuge at 8000rpm for 1 minute, recover the supernatant for later use.

[0052] Ligation of recovered fragments to the vector:

[0053]

[0054] Ligation at 14°C for 18 hours.

Embodiment 3

[0055] Embodiment 3, recombinant transformation Escherichia coli DH5α bacterial strain:

[0056] Take 5 ul of the ligation product, add 100 ul of competent bacteria, mix well, ice bath for 30 minutes, heat shock at 42°C for 1 minute, ice bath for 2 minutes, add 800ul LB medium, and incubate at 37°C for 1 hour. Take 100 ul of the bacterial solution and spread it on a plate containing ampicillin, and incubate it upside down at 37°C for 16 hours.

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Abstract

The invention relates to a gland virus carrier for expressing Math1 albumen and a transformant thereof, and a construction method of the carrier. The gland virus for expressing Math1 albumen obtained by the method can overcome one or a plurality of defects of the existing medication route and is safely and effectively used for treating sensorineural hearing loss.

Description

technical field [0001] The invention belongs to the field of gene therapy for sensorineural deafness, and specifically relates to the construction of a recombinant adenoviral vector for inner ear-specific expression gene Math1, intracochlear transfection and its application in gene therapy for sensorineural deafness. Background technique [0002] Cochlear hair cells are terminally differentiated cells. As mechanosensory cells, they can convert sound stimuli into electrical signals and play a vital role in maintaining hearing. Aging, ototoxic drugs, infection, heredity, excessive acoustic stimulation and autoimmune diseases can cause irreversible damage to cochlear hair cells, leading to permanent sensorineural deafness. Sensorineural deafness affects tens of millions of people around the world. With the acceleration of human aging and industrialization, this problem has gradually become prominent and has become an important issue in the field of public health. [0003] Alth...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/861C12N7/01C12N15/12A61K48/00A61P27/16
Inventor 杨仕明翟所强高维强郭维维胡吟燕陈伟申卫东王秋菊韩东一杨伟炎
Owner 杨仕明
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