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Recombinant plasmid, recombinant virus vector, recombinant virus strain and application thereof, recombinant protein and subunit vaccine containing the same

A recombinant viral vector, recombinant virus technology, applied in the direction of virus/phage, application, viral peptide, etc., can solve the problems of short protective effect, enhanced antibody dependence, prone to internal errors, etc., and achieve good immunogenicity.

Inactive Publication Date: 2015-11-11
JILIN HEYUAN BIOENG LIMITED
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] As an RNA virus, the gene of PRRSV is prone to internal errors during synthesis, the ability of inactivated vaccines to induce cellular immunity is weak, and the protective effect is short. PRRSV also has "antibody-dependent enhancement"
Therefore, there are certain drawbacks in the development of inactivated vaccines and attenuated vaccines for preventing PRRS.

Method used

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  • Recombinant plasmid, recombinant virus vector, recombinant virus strain and application thereof, recombinant protein and subunit vaccine containing the same
  • Recombinant plasmid, recombinant virus vector, recombinant virus strain and application thereof, recombinant protein and subunit vaccine containing the same
  • Recombinant plasmid, recombinant virus vector, recombinant virus strain and application thereof, recombinant protein and subunit vaccine containing the same

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Experimental program
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Embodiment 1

[0039] Embodiment 1: Construction of recombinant baculovirus (BacSC-Dual-GP5) of the present invention and expression of recombinant protein

[0040] 1 Materials and methods

[0041] 1.1 Viruses, cells and antiserum

[0042] Porcine Reproductive and Respiratory Syndrome Virus Strain: It was presented by Professor Ding Zhuang from the Department of Infectious Diseases, School of Veterinary Medicine, Jilin University. The cell culture adopts DMEM medium, supplemented with 8-12% fetal bovine serum, 100 U / ml penicillin and 100 μg / ml streptomycin. Carry out cell culture by routine method, inoculate virus when Marc-145 cell grows to culture surface monolayer 75-80%, cell subculture effect is good (Lan Hainan, Zhao Xiaoli, etal. pig blue ear disease CH-IR strain (PRRSV- CH-IR) cultured in Hou kidney cells (marc-145) [J]. Zoonotic Diseases and Veterinary Public Health, 2008.231-234). Grace's insect cell culture medium and insect sf-9 cells were used to culture the baculovirus vector ...

Embodiment 2

[0056] Example 2 Preparation of subunit vaccine with the PRRSV-GP5 protein expressed by the recombinant baculovirus strain (BacSC-Dual-GP5) constructed by the present invention

[0057] The recombinant baculovirus strain (BacSC-Dual-GP5) constructed by the present invention is used to inoculate the PRRSV-GP5 protein product expressed by insect cells as an antigen to prepare a subunit vaccine. The research experiment optimized the expression parameters of the recombinant protein in insect cells, inoculated the insect cells with 5 infection units, and used the expressed 5 batches of recombinant protein as antigens to develop a subunit vaccine. The recombinant protein culture and Preparation of subunit vaccine by emulsification of immune adjuvant. The semi-finished and finished vaccines were inspected according to the requirements of the veterinary biological products regulations, and the clinical trials were carried out with the qualified products.

[0058] Test results: Five 2...

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Abstract

The invention relates to the technical field of animal genetic engineering and animal virology and especially relates to a recombinant plasmid, a recombinant virus vector, a recombinant virus strain and an application thereof, a recombinant protein and a subunit vaccine containing the same. In the invention, a recombinant baculovirus strain BacSC-Dual-GP5, which high-effectively expresses porcine reproductive and respiratory syndrome virus GP5 protein, is constructed, thereby achieving a better immunogenicity. The recombinant protein expressed by the recombinant virus strain is free of a fusion part and is very high in product expression yield. The PRRSV-GP5 subunit vaccine product in the invention is safe to animal. An immune efficacy test result of the vaccine proves that after 28 days that all vaccines are subjected to immunization, all antibody detection shows positive. A strong poison challenge test proves that both a 1 ml immune group and a 2 ml immune group achieve 100% protection while a 0.5 ml group achieves 80% (4 / 5) protection.

Description

technical field [0001] The invention relates to the technical fields of animal genetic engineering and animal virology, in particular to recombinant plasmids, recombinant virus vectors, recombinant virus strains and applications thereof, recombinant proteins and subunit vaccines containing the proteins. Background technique [0002] Porcine reproductive and respiratory syndrome (PRRS) is a viral disease that seriously harms pig herds. The main features are reproductive failure in sows and dyspnea in finishing pigs and piglets. GP5 is an important glycoprotein of PRRSV, which can induce neutralizing antibody production and immune protection. Therefore, it is the main target site for the study of genetically engineered vaccines against PRRS. Although inactivated and modified live vaccines are available, their protection is no longer complete. Therefore, in order to study an effective vaccine against PRRSV infection, a new technology must be developed. [0003] As an RNA vi...

Claims

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Application Information

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IPC IPC(8): C12N15/63C12N15/866C12N7/01C07K14/08A61K39/12A61P31/14
Inventor 李秋菊
Owner JILIN HEYUAN BIOENG LIMITED
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