Multivalent adenovirus vaccine for preventing toxoplasmosis

A technology for toxoplasmosis and adenovirus, applied in the field of adenovirus vaccines, can solve problems such as the difficulty of vaccine protection, and achieve the effects of improving immune response and survival rate, reducing the formation of cysts in the brain, and prolonging survival time.

Active Publication Date: 2014-10-29
SHANDONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, it is difficult to form a comprehensive and effective protective effect for vaccines that only target antigens in a certain period of time.

Method used

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  • Multivalent adenovirus vaccine for preventing toxoplasmosis
  • Multivalent adenovirus vaccine for preventing toxoplasmosis
  • Multivalent adenovirus vaccine for preventing toxoplasmosis

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Example 1: Construction of Toxoplasma gondii polyvalent adenovirus vaccine

[0029] Such as figure 1 As shown, first from the Toxoplasma gondii tachyzoite antigens SAG3, ROP18, MIC6, GRA7, bradyzoite antigens MAG1, BAG1 and sporozoite antigen SPA genes, IEDB software was used to screen out epitopes that are concentrated and have both human and BALB / C mouse MHC molecularly restricted gene segment: SAG3 101-144 ,ROP18 347-396 ,MIC6 288-347 ,GRA7 182-224 ,MAG158-125 ,BAG1 156-211 ,SPA 142-200 and a proven effective CD4 + The T cell epitope AS15 gene fragment constitutes a linear compound multivalent antigen gene, and its N-terminus is linked to the ubiquitin protein gene, which is synthesized by Suzhou Jinweizhi Biotechnology Co., Ltd.

[0030] Among them, SAG3 101-144 The nucleotide sequence of the gene fragment is: ggcttgggcg gagagttttt gccgctcgaa ggcggcacgt cgtcgtaccc gcgagtatgt cacattgatg ccaaggacaa gggcgactgc gagcgcaaca agggctttct gaccgactac ataccgggcg cg, as...

Embodiment 2

[0041] Embodiment 2: vaccine immunization BALB / c mice

[0042] SPF grade female BALB / c mice (6-8 weeks) were purchased from the Experimental Animal Center of Shandong University. 40 mice were randomly divided into 6 groups, and each group of mice in the experimental group was intramuscularly injected with 3x10 8PFU, the adenovirus vaccine Ad-UbTgMEG that the embodiment of the present invention 1 prepares, the control group injects immunization 3x10 8 PFU empty virus Ad-GFP and 100 μl PBS. Mice were immunized twice with an interval of three weeks.

Embodiment 3

[0043] Embodiment 3: the mensuration of immune mouse humoral immunity level

[0044] On the 0th, 14th, 35th, and 49th days, the blood was collected from the inner canthus of the mice. The blood samples were first allowed to stand at room temperature for 3 hours, and then centrifuged at 3000 rpm for 30 minutes to collect serum. Serum levels of total anti-T. gondii immunoglobulins (IgG1 and IgG2a) were determined using enzyme-linked immunosorbent assay. The result is as Figure 3a to Figure 3b shown. After two immunizations, compared with the control group, PBS and Ad-GFP groups, the antibody titer level of the vaccine immunization group increased significantly. On the 35th day and 49th day, the mice in the Ad-UbTg. High levels of Toxoplasma-specific IgG antibodies were detected (P<0.01).

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Abstract

The invention discloses a compound multivalent antigen gene, a recombinant plasmid and an adenovirus vaccine for preventing toxoplasmosis. The preparation process comprises the following steps: connecting a ubiquitin protein gene, a toxoplasmosis tachyzoite antigen gene, a bradyzoite antigen gene, a sporozoite antigen SPA gene and an AS15 gene, and synthetizing the compound multivalent antigen gene for preventing toxoplasmosis; carrying out double digestion on the synthesized target gene and then inserting the synthesized target gene between BamHI and SmaI of a vector to generate the recombinant plasmid; and carrying out digestion, PCR and sequencing identification on the recombinant plasmid, extracting plasmid and purifying; transferring the recombinant plasmid and skeleton plasmid into cells together, packing and removing toxins, so as to obtain the recombinant adenovirus vaccine. By adopting the adenovirus vaccine disclosed by the invention, toxoplasma infection of people or animals can be effectively prevented, formation of brain tissue cysts in the chronic infection process of toxoplasma is controlled, the immunologic reaction and the survival rate are improved in acute infection, and candidate components of an effective vaccine with comprehensive protection are provided.

Description

technical field [0001] The present invention relates to an adenovirus vaccine for preventing toxoplasmosis, in particular to an adenovirus vaccine based on the connection of protective antigen fragments of tachyzoites, bradyzoites and sporozoites of toxoplasma gondii Multi-antigen compound gene, and ubiquitin protein gene as adjuvant, construct recombinant adenovirus as carrier of toxoplasma polyvalent adenovirus vaccine. Background technique [0002] Toxoplasma gondii (Toxoplasma gondii) is a worldwide distribution of parasitic protozoa, widely parasitic in nucleated cells of humans and animals, causing serious zoonotic parasitic diseases. The general population in my country is generally susceptible to Toxoplasma gondii, with an average infection rate of about 7.89%. For special populations (such as pregnant women), the infection rate can be as high as 10%. Infection in early pregnancy women can lead to miscarriage, premature birth or even stillbirth, and survivors often ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K48/00A61K39/235A61P33/02
Inventor 丛华袁泉赵玲潇丛海滋张晓丽尹绘权
Owner SHANDONG UNIV
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