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Pseudorabies virus gene deleted strain, inactivated vaccine for porcine pseudorabies, and preparation method and application of inactivated vaccine

A technology of pseudorabies virus and porcine pseudorabies, which is applied in the field of bioengineering, can solve the problems of poor protection effect of mutant strains, and achieve the effects of reducing cost burden, long immunization cycle and good safety

Pending Publication Date: 2020-02-28
SHANGHAI ACAD OF AGRI SCI +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0009] The present invention aims to solve the technical problem of PRV mutant strains appearing in China at present, but the existing PRV vaccines have poor protection effect on the mutant strains, and it is urgent to develop new PRV vaccines, and provides a gene deletion strain of pseudorabies virus, which is composed of The gE gene deletion strain constructed by the popular strain of pseudorabies virus, the porcine pseudorabies inactivated vaccine prepared by using the gene deletion strain has good immunogenicity and significantly enhanced immune effect

Method used

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  • Pseudorabies virus gene deleted strain, inactivated vaccine for porcine pseudorabies, and preparation method and application of inactivated vaccine
  • Pseudorabies virus gene deleted strain, inactivated vaccine for porcine pseudorabies, and preparation method and application of inactivated vaccine
  • Pseudorabies virus gene deleted strain, inactivated vaccine for porcine pseudorabies, and preparation method and application of inactivated vaccine

Examples

Experimental program
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Effect test

Embodiment 2

[0082] The preparation of embodiment 2 porcine pseudorabies inactivated vaccine

[0083] The preparation method of porcine pseudorabies inactivated vaccine comprises the following steps:

[0084] (1) Pig testicular cells are cultured in a bioreactor for suspension culture, the temperature of the suspension culture is controlled at 37°C, the DO is 50%, the stirring speed is 100r / min, and the pH is 7.2;

[0085] (2) When the cell density is 3*10 6 When inoculating the pseudorabies virus gE gene deletion strain constructed in Example 1, its virulence TCID 50 ≥10 8.75 / ml, the virus strain is diluted to a concentration of 0.1% by weight;

[0086](3) When more than 80%-90% of the above-mentioned pig testicular cells have lesions, harvest the cell culture, freeze and thaw repeatedly 3 times, and obtain the cell venom containing the supernatant; store the cell venom below -20°C , the retention period shall not exceed 30 days;

[0087] (4) Take the supernatant to measure the pois...

Embodiment 3

[0096] The preparation of embodiment 3 porcine pseudorabies inactivated vaccine

[0097] The preparation method of porcine pseudorabies inactivated vaccine comprises the following steps:

[0098] (1) Pig testicular cells are cultured in a bioreactor for suspension culture, the temperature of the suspension culture is controlled at 37°C, the DO is 50%, the stirring speed is 100r / min, and the pH is 7.2;

[0099] (2) The pseudorabies virus gE gene deletion strain constructed in Example 1, its virulence TCID 50 ≥10 8.75 / ml, the virus strain is diluted to a weight percent concentration of 0.1%, and the density obtained by inoculating to step (1) is 3*10 6 In the suspended porcine testicular cells, the temperature is 37°C, the DO is 50%, the stirring speed is 90r / min, and the pH is 7.2;

[0100] (3) When more than 80%-90% of the cells have cytopathic changes, harvest the cell culture, freeze and thaw repeatedly 3 times to obtain the cell venom containing the supernatant, and sto...

Embodiment 4

[0104] The preparation of embodiment 4 porcine pseudorabies inactivated vaccine

[0105] The preparation method of porcine pseudorabies inactivated vaccine comprises the following steps:

[0106] (1) Pig testicular cells are cultured in a bioreactor for suspension culture, the suspension culture control temperature is 35°C, DO is 40%, stirring speed is 80r / min, and pH is 7.2;

[0107] (2) The pseudorabies virus gE gene deletion strain constructed in Example 1, its virulence TCID 50 ≥10 8.75 / ml, the virus strain is diluted to a weight percent concentration of 0.1%, and the density obtained by inoculating to step (1) is 3*10 6 In the suspended porcine testicular cells, the temperature is 35°C, the DO is 40%, the stirring speed is 80r / min, and the pH is 7.2;

[0108] (3) When more than 80%-90% of the cells have cytopathic changes, harvest the cell culture, freeze and thaw repeatedly 3 times to obtain the cell venom containing the supernatant, and store the cell venom below -2...

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Abstract

The invention discloses a pseudorabies virus gene deleted strain, and a preparation method of an inactivated vaccine for porcine pseudorabies. The gene deleted strain is a pseudorabies virus gE gene deleted strain, which is constructed through reading frame frameshift mutation caused by deletion of a plurality of basic groups from a wild-type pseudorabies virus (PRV) strain gE gene sequence. The preparation method includes the following steps: constructing the pseudorabies virus gE gene deleted strain; domesticating a porcine testicular cell and subjecting the same to suspension culture; inoculating the porcine testicular cell with the pseudorabies virus gE gene deleted strain, and harvesting a cell culture when 80-90% of the cell has lesions to obtain cell venom containing supernatant; taking the supernatant to measure the valence, and inactivating the qualified cell venom in a sterilization container; and mixing the inactivated cell venom with an adjuvant to obtain the inactivated vaccine for the porcine pseudorabies. The prepared inactivated vaccine for the porcine pseudorabies has high immunogenicity, long immunization period, and no side effects after immunization, is safe andreliable, and can effectively prevent infections of pseudorabies virus epidemic strains.

Description

technical field [0001] The invention relates to the technical field of bioengineering, in particular to a gene-deleted strain of pseudorabies virus, an inactivated porcine pseudorabies vaccine and a preparation method and application thereof. Background technique [0002] Pseudorabies (Pseudo rabies, PR) is a severe infectious disease caused by pseudorabies virus (Pseudorabies virus, PRV) that occurs in domestic animals and wild mammals, with fever, itching (except pigs) and encephalomyelitis as the main symptoms. The disease is a typical natural foci disease, pigs are its main host and source of infection, and the economic losses caused to the pig industry are second only to swine fever. In other animals except pigs, the disease mostly occurs in a sporadic form, and after the disease occurs, it all ends in death. [0003] Due to the loss caused by pseudorabies is second only to foot-and-mouth disease and swine fever, it has become one of the swine diseases that some Europe...

Claims

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Application Information

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IPC IPC(8): C12N7/01A61K39/245A61P31/22G01N33/569
CPCC12N7/00C07K14/005A61K39/12A61P31/22G01N33/56994C12N2710/16721C12N2710/16722C12N2710/16734A61K2039/552A61K2039/5252G01N2333/032
Inventor 彭丽英林鸷何錫忠
Owner SHANGHAI ACAD OF AGRI SCI
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