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Vaccine for preventing herpesvirus hominis type II

A herpes simplex virus, vaccine technology, applied in antiviral agents, medical preparations containing active ingredients, gene therapy and other directions, can solve the problems of affecting practical application, low efficiency, etc. The effect of morbidity

Inactive Publication Date: 2015-06-24
ZHEJIANG ACAD OF MEDICAL SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

DNA vaccines can induce humoral immunity and cellular immunity in the immune response, and have been successfully applied to various animals such as mice, sheep, chickens, monkeys and chimpanzees, but their efficiency in inducing immune responses is relatively low , affecting its practical application, so finding molecules that can improve the immune efficiency of nucleic acid vaccines is an urgent problem to be solved

Method used

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  • Vaccine for preventing herpesvirus hominis type II
  • Vaccine for preventing herpesvirus hominis type II
  • Vaccine for preventing herpesvirus hominis type II

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0016] Embodiment 1: the acquisition of eukaryotic expression plasmid

[0017] The eukaryotic expression plasmid vector pcDNA3-Kan was constructed by replacing the original ampicillin resistance (Amp) on the vector pcDNA3 with Kanna resistance (Kan) by molecular biology techniques PCR, enzyme digestion, ligation and clone screening. The complete gene sequence (gD) of glycoprotein D of HSV-2 sav strain was amplified by PCR method, and the recombinant plasmid pcDNA-Kan / gD containing gD was obtained after cloning and recombination. The recombinant plasmid pcDNA-Kan / gD and its specific preparation method have been compared with "He Fang et al. Chinese Journal of Health Inspection, 2008, 18(6): 997-999".

Embodiment 2

[0018] Embodiment 2: the acquisition of recombinant plasmid

[0019] The activated T cells were taken, the total RNA was extracted with RNeasy Mini Kit (QIAGEN), and the first-strand cDNA was synthesized by reverse transcription with Oligod(T)n as a primer. Referring to the sequence number BC119225.1 included in the NCBI nucleic acid database, a pair of primers were designed, and the synthesized cDNA was used as a template to amplify to obtain SEQ ID NO.1. The upstream primer sequence is 5 ， -GCC GAATTC ATGATAGAAACATACAGCCAACCT-3 ， , the downstream primer sequence is 5 ， -GAC CTCGAG TCAGAGTTTGAGTAAGCCAAAAGA-3 ， .

[0020] The gene sequence shown in SEQ ID NO.1 and pcDNA3-Kan were digested with EcoRI and Xho I, purified and recovered by gel cutting. The resulting fragments were recovered and ligated with T4 ligase at a molar concentration of 1:5. The ligation product was transformed into a competent strain of DH5α, and the next day, a medium-sized white colony was pic...

Embodiment 3

[0022] Embodiment 3: The plasmid that embodiment 1 and 2 obtain is used for immune mouse

[0023] 1. Experimental materials:

[0024] 1) The plasmids obtained in Examples 1 and 2 were respectively amplified in E.coli and purified with QIAGEN purification kit. The concentration and purity of the plasmid were measured by ultraviolet spectrophotometry. The concentration and purity of the DNA were determined by OD260 and OD280. The plasmid DNA with a ratio of OD260 / OD280 of 1.8-2.0 was selected to immunize mice. The two plasmids were cryopreserved at -20°C, thawed before immunizing mice, and the two plasmids were mixed according to the mass ratio of 2:1.

[0025] 2) Frozen HSV-2 virus sav3 was repeatedly frozen and thawed three times. After centrifugation, the supernatant was added to the monolayer of Vero cells and incubated at 37°C for 1 h. Remove the virus liquid, add 4% FBS DMEM complete medium, and incubate at 35°C for 24-48h. Adherent cells were scraped off with a cell s...

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Abstract

The invention discloses a vaccine for preventing herpesvirus hominis type II. The glycoprotein D gene of the herpesvirus hominis type II is combined into a eukaryotic expression vector pKan with Kanamycin resistance by a molecular biology method, so as to construct a DNA vaccine containing a eukaryotic expression plasmid pgD containing a full-length glycoprotein D gene and a recombinant plasmid containing a gene sequence as shown in SEQ ID NO.1. The body immunized with the DNA vaccine can produce effective humoral immunity and cellular immunity to prevent and treat diseases caused by infection with the herpesvirus hominis type II, for example, herpes progenitalis.

Description

technical field [0001] The invention belongs to a biological product, in particular to a vaccine for preventing herpes simplex virus type II (HSV-2). Background technique [0002] Genital herpes is a sexually transmitted disease that is relatively common in the world and in coastal areas of my country, and its incidence is gradually increasing. It is caused by the human herpes simplex virus (HSV) that infects the skin and mucous membranes of the genitourinary and anus through sexual transmission. Herpes simplex virus can be divided into types 1 and 2. Most genital herpes is caused by HSV-2 infection. The main source of infection is genital herpes patients and asymptomatic virus carriers. HSV-2 infection is lifelong, the virus has neurotropic latency, and the virus can establish lifelong latency in the human sacrococcygeal ganglion. HSV-2 latent infection can lead to recurrence of relevant clinical symptoms due to activation of various non-specific stimuli, such as typical g...

Claims

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Application Information

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IPC IPC(8): A61K48/00A61K39/245A61P31/22
Inventor 陶薇洪艳胡如西傅婷何卓晶贾岚
Owner ZHEJIANG ACAD OF MEDICAL SCI
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