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Preparation and application of recombinant adenovirus capable of expressing IL-10

A recombinant adenovirus and adenovirus technology, applied in the field of biomedical genetic engineering, can solve the problems of long action time and short protein action time, and achieve the effects of less injection volume, strong pertinence and high transfection efficiency

Inactive Publication Date: 2018-01-30
UNIV OF ELECTRONIC SCI & TECH OF CHINA
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  • Application Information

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Problems solved by technology

The IL-10 protein has a certain effect in the short term, which shows that IL-10 has an anti-inflammatory effect, but the protein's action time is short, and after a week, it is metabolized by the mouse and no longer works, especially for In chronic mild stress (CMS) depression-like model mice, the effect is not achieved. Therefore, there is an urgent need for a method that expresses endogenous proteins and acts for a long time

Method used

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  • Preparation and application of recombinant adenovirus capable of expressing IL-10
  • Preparation and application of recombinant adenovirus capable of expressing IL-10
  • Preparation and application of recombinant adenovirus capable of expressing IL-10

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Experimental program
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Effect test

Embodiment Construction

[0048] 1. Construction of pAAV2 / 5-GFAP-IL10 vector

[0049] 1.1 Cloning the target gene

[0050] Extraction of total RNA from mouse spleen: C57 / 6J mice were sacrificed by cervical dislocation, immersed in 75% alcohol, fixed on a splint after 5 minutes, the spleen was taken out, and RNA was extracted under liquid nitrogen conditions; the cloning of the target gene: using the extracted RNA , synthesize DNA by reverse transcription, use DNA as a template, and use primers designed upstream and downstream (including BamH I and Xho I restriction sites in the primers) to carry out PCR amplification (the reaction conditions for PCR amplification are: 94°C 5min; 94°C ℃ 30sec, 55℃ 30sec, 72℃ 60sec, a total of 34 cycles; stored at 4℃), the amplified product was identified correctly by 1% agarose gel electrophoresis, and the gel was cut and recovered, and a DNA fragment with a size of about 537bp was recovered.

[0051] 1.2 Construction of the carrier

[0052] Construction of plasmid pA...

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Abstract

The invention provides a preparation method and application of adeno-associated virus capable specifically over-expressing interleukin 10 in astrocyte. The virus is a recombinant adenovirus which is established by replacing a broad spectrum promoter (CMV) into a specific promoter (GFAP) of the astrocyte on a type 2 adenovirus vector and inserting a mIL-10 gene at the MCS position before T2A. The recombinant adenovirus can be applied to preparation of anti-depression medicines.

Description

technical field [0001] The invention relates to the field of biomedical genetic engineering, in particular to the preparation and application of a recombinant adenovirus expressing IL-10. Background technique [0002] Depression has become a mental disease that seriously endangers human physical and mental health. According to the statistics of the World Health Organization, about 350 million people around the world suffer from depression, which has become the fourth largest disease in the world. Depression is a mental disorder characterized by significant and persistent depression. Its main manifestations are depression, slow thinking and psychomotor inhibition. At present, the treatment of depression mainly relies on psychotherapy and adjuvant drug therapy. There are huge potential business opportunities in the antidepressant drug market, but due to the side effects and tolerance of antidepressant drugs, and poor pertinence. There is an urgent need for close cooperatio...

Claims

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Application Information

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IPC IPC(8): C12N7/01C12N15/66C12N15/70A61K38/20A61K48/00A61P25/24
Inventor 游自立张立娟
Owner UNIV OF ELECTRONIC SCI & TECH OF CHINA
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