DNA vaccine for preventing and treating cysticercosis cellulosae and preparation method thereof

A DNA vaccine, a technology for swine cysticercosis, which can be used in gene therapy, pharmaceutical formulations, genetic material components, etc., can solve the problems of poor cell line stability, large side effects, and unsatisfactory curative effects, and achieve stable physical and chemical properties and low cost. , the effect of good immune protection

Inactive Publication Date: 2009-09-09
SECOND MILITARY MEDICAL UNIV OF THE PEOPLES LIBERATION ARMY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] At present, most of the treatment options for cysticercosis are conventional drug therapy, the curative effect is not ideal, and the side effects are relatively large, and it can only be used for post-infection treatment rather than prevention
Vaccine is an important means to prevent cysticercosis. At present, the research on cysticercosis vaccine in China mainly has two aspects: one is the development of porcine cysticercosis cell vaccine. The cell vaccine has good immunity, but the stability of the cell line is relatively low. Poor; the other is a DNA vaccine

Method used

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  • DNA vaccine for preventing and treating cysticercosis cellulosae and preparation method thereof
  • DNA vaccine for preventing and treating cysticercosis cellulosae and preparation method thereof
  • DNA vaccine for preventing and treating cysticercosis cellulosae and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Example 1: Cloning of the Anx B2 gene from Hexacarpa

[0037]1. Activation of Taenia solium, extraction of total RNA and RT-PCR, PCR amplification

[0038] Fresh tapeworm proglottids collected from Taenia solium patients (purchased from Harbin Medical University) were rinsed several times with saline, cut into pieces in a 7ml Eppendorf tube and squeezed to release the eggs, centrifuged and rinsed 3 times at 3000r / min ; Add an equal amount of sodium hypochlorite to act for a few minutes (microscopic examination of most eggs broken), quickly add an equal amount of 0.2mol / L sodium thiosulfate to terminate the reaction, centrifuge at 3000r / min to remove egg fragments; resuspend with normal saline Precipitate, add hatching activation solution containing trypsin and pig bile, incubate at 37°C for 40 minutes (violently shake once every 10 minutes), centrifuge to discard the supernatant, and the precipitate is the activated Hexacarpa. Weigh 30 mg of Hexacarpa precipitate and p...

Embodiment 2

[0045] Example 2: Construction and preparation of pCDNA3.1-Anx B2 vaccine

[0046] 1. Construction of pCDNA3.1-Anx B2 DNA vaccine

[0047] Using the plasmid pGEM-T-Anx B2 containing the target gene as a template, PCR amplification was carried out with the above-mentioned primers 1 and 2. After the PCR product was recovered from the gel, it was double-digested with BamH I and xho1 (purchased from NEB Company), and the same BamH I and xho1 double-digested pCDNA3.1 vector (purchased from Invitrogen) was connected by T4 ligase, transformed into Escherichia coli DH5α, positive recombinants were obtained by blue-white screening, extracted plasmid, BamH I, xho1 digestion and sequencing identification, sequencing The result was correct, and the engineering bacteria DH5α containing the recombinant plasmid pCDNA3.1-Anx B2 was obtained. (For specific experimental steps, refer to the third edition of "Molecular Cloning", Science Press, published in 2002 for related content)

[0048] 2. ...

Embodiment 3

[0051] Example 3: In vitro expression of pCDNA3.1-Anx B2 vaccine

[0052] The constructed DNA vaccine was transfected into COS7 cells by electroporation (see the third edition of "Molecular Cloning" for specific steps, published by Science Press in 2002), and the cells were harvested after 72 hours, and detected by indirect ELISA. The expression level of the antigen Anx B2 in the supernatant and cell lysate was positive after color development, and the P / N ratio was greater than 1.8. The result was positive in the supernatant and cell lysate, and the AnxB2 antigen was expressed in COS7 cells. It shows that the DNA vaccine can be expressed in mammalian cells.

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Abstract

The invention relates to the technical field of biology, in particular to a DNA vaccine for preventing and treating cysticercosis cellulosae and a preparation method thereof. The prior therapeutic schedule for treating the cysticercosis cellulosae adopts common medicine, has unfavorable curative effect, large side effect and can be only used for treating after the infection but not prevention. The DNA vaccine for preventing and treating cysticercosis cellulosae comprises eukaryotic expression vector pcDNA3.1 and cysticercosis cellulosae antigen gene Anx B2. The preparation method includes the step that the cysticercosis cellulosae antigen gene Anx B2 is cloned from oncosphere. The invention can effectively induce the immune response reaction of piglets, has a certain immune protection function to animals infecting the cysticercosis cellulosae, and has simple production technology, low cost, stable physicochemical property and convenient storage and transportation.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a DNA vaccine for preventing and treating porcine cysticercosis and a preparation method thereof. Background technique [0002] Cysticercosis, short for cysticercosis, is a zoonotic disease caused by infecting pigs or humans with the larvae of Taenia solium - cysticercosis. The disease is widely prevalent all over the world, and it is more common in developing countries such as Brazil and India, which seriously threatens human health and animal husbandry production. China is one of the endemic countries of cysticercosis. [0003] At present, most of the treatment options for cysticercosis are conventional drug therapy, which has unsatisfactory curative effect and relatively large side effects, and can only be used for post-infection treatment rather than prevention. Vaccine is an important means to prevent cysticercosis. At present, the research on cysticercosis vaccine in China mai...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K48/00A61P33/14
Inventor 王凯慧孙树汉程娜陆一鸣王俊杰张毅王娜魏玉保
Owner SECOND MILITARY MEDICAL UNIV OF THE PEOPLES LIBERATION ARMY
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