HBV-infected mouse model construction method and application

A construction method and mouse model technology, applied in the field of medicine, can solve the problems of inability to produce cccDNA, limited duration, etc., and achieve the effect of simple preparation method and simple establishment method

Active Publication Date: 2016-09-21
SECOND MILITARY MEDICAL UNIV OF THE PEOPLES LIBERATION ARMY
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Problems solved by technology

Although both DHBV and WHV can cause chronic infection, they are different from human HBV in gene structure and pathogenicity; the HBV transgenic mouse model integrates the HBV genome in the chromosome, is immune to HBV, and cannot produce cccDNA; hig

Method used

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  • HBV-infected mouse model construction method and application
  • HBV-infected mouse model construction method and application
  • HBV-infected mouse model construction method and application

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Embodiment 1

[0022] 1. Materials

[0023] 1. Cells and plasmids

[0024] Human liver cancer cells Huh7, HepG2, and human embryonic kidney (HEK) 293T cell lines were preserved by the Department of Microbiology, Second Military Medical University (or directly purchased from the Institute of Cells, Chinese Academy of Sciences); the pMD-18T cloning vector and plasmid pUC18 are Takara products.

[0025] 2. Plasmid

[0026] The adeno-associated virus backbone plasmid pAAV-HBV1.2 containing 1.2 copies of the HBV genome was donated by Professor Chen Peizhe of National Taiwan University (it can also be prepared according to the following literature methods: Huang LR, Wu HL, ChenPJ, Chen DS. An immunocompetent mouse model for the tolerance of human chronic hepatitis B virus infection. Proc Natl Acad Sci US A. 2006; 103(47): 17862-7.); gene cloning vector pMD-18T is a product of Takara Company.

[0027] 3. Primers

[0028] The primers for amplifying the complete HBV genome were synthesized by Taka...

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Abstract

The invention relates to the technical field of medicine and provides a hepatitis B virus (HBV)-infected mouse model construction method. According to the method, a polymerase chain reaction (PCR) is adopted to amplify an HBV single-copy linear genome, a PCR product transfects Huh7 cells, HepG2 cells and 293T cells, and hepatitis b surface antigen (HBsAg) and hepatitis B e antigen (HBeAg) in supernatant of culture cells are detected through an enzyme-linked immunosorbent assay (ELISA); the PCR product is injected to a C57BL/6J mouse through a tail vein high-pressure hydrodynamic method, so that hepatitis b core antigen (HBcAG) in mouse liver tissue is detected in an immunohistochemistry mode, and the HBsAg and the HBeAG in serum of the mouse are detected in an ELISA mode. By means of the method, a novel HBV-infected mouse model is constructed, the model is simple in preparation method, and the HBC genome similar to the HBV cccDNA exists in the liver tissue. A novel tool is provided for research on an HBV cccDNA degradation mechanism, medicine removal, HBV biological characteristics in a serum sample of a hepatitis B patient and the like.

Description

Technical field: [0001] The invention relates to the technical field of medicine, in particular to a mouse model of HBV infection established by injecting HBV genome PCR products into mice, and the application of the model. Background technique: [0002] Hepatitis B virus (HBV) is a pathogen that seriously endangers human health. At present, about 300 million people in the world are chronically infected with HBV. HBV infection can not only cause acute and chronic hepatitis, but also is an important cause of liver cirrhosis and liver cancer in developing countries. Although hepatitis B vaccination can effectively prevent HBV infection, for chronic HBV infection, current drugs can only inhibit virus replication, but cannot act on HBV covalently closed circular DNA (cccDNA) present in the nucleus of liver cells, which is an anti-viral agent. Huge challenges for HBV drug development (see literature: Baltayiannis G, Karayiannis P. Treatment options beyond IFNα and NUCs for chron...

Claims

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Application Information

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IPC IPC(8): A61K35/74A61K49/00
CPCA61K35/74A61K49/0008
Inventor 赵平张龙严狄弘玮王世杰张卫
Owner SECOND MILITARY MEDICAL UNIV OF THE PEOPLES LIBERATION ARMY
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