Porcine pseudorabies virus gene deletion strain, vaccine composition and its preparation method and application

A technique for porcine pseudorabies virus and vaccine composition, applied in the field of animal virology

Active Publication Date: 2019-01-15
PULIKE BIOLOGICAL ENG INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Pigs immunized with vaccines in the prior art cannot completely resist the wild virus attack, and symptoms such as high fever, depression, loss of appetite or extinction will still occur, the infection rate exceeds 80%, the morbidity rate exceeds 30%, and the mortality rate is between 10% and 10%. Between 20%
For example, Peng Jinmei et al. Isolation and identification of new epidemic strains of porcine pseudorabies virus and analysis of antigenic differences. Chinese Journal of Preventive Veterinary Medicine, 2013, 35(1): 1-4; Tong Wu et al. Isolation of pseudorabies virus in infected piglets after immunization and Identification. Chinese Journal of Animal Infectious Diseases, 2013,21(3):1-7; Yu et al., Pathogenic Pseudorabies Virus, China, 2012.Emerging infectious Diseases.2014,20(1):102-104; An et al ., Pseudorabies virus variant in Bartha-K61-vaccinated pigs, China, Emerging infectious Diseases.2013.19 (11): 1749-1755), there is no vaccine in the prior art to solve the pseudorabies caused by the pig pseudorabies variant

Method used

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  • Porcine pseudorabies virus gene deletion strain, vaccine composition and its preparation method and application
  • Porcine pseudorabies virus gene deletion strain, vaccine composition and its preparation method and application
  • Porcine pseudorabies virus gene deletion strain, vaccine composition and its preparation method and application

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0074] Embodiment 1, the preparation of PRV HN1201 strain RR deletion strain

[0075] 1.1 Construction of GFP intermediate transfer vector required for PRV HN1201RR deletion

[0076] According to the sequence SEQ ID NO.2 of the RR gene to be deleted, design homology arms at its two ends, respectively RRA and RRB. RRA and RRB were cloned into pUC19 vector and named pUCRRAB. Then the GFP gene was cloned between RRA and RRB on pUCRRAB to obtain a recombinant virus transfer vector, named pUCRRA-GFP-B. The homologous arms in the transfer vector are the sequences on both sides of RR, so the recombinant virus obtained after recombination is the deletion of RR gene. The schematic diagram of recombinant transfer vector construction is shown in figure 1 , figure 2 is the location of the RRA and RRB homology arms on the genome.

[0077] 1.1.1. Amplification and cloning of homologous recombination arm

[0078] 1.1.1.1 Primer design and template preparation

[0079] According to th...

Embodiment 2

[0120] Embodiment 2, the preparation of PRV HN1201 strain TK / gE / gI / deletion strain

[0121] 1. Preparation of TK deletion strain

[0122] 1.1 Construction of GFP intermediate transfer vector required for deletion of PRV HN1201TK

[0123] According to the sequence of the TK gene to be deleted, design homology arms at its two ends, respectively TKA and TKB. TKA and TKB were cloned into pUC19 vector, named pUCTKAB. Then the GFP gene was cloned into pUCTKAB to obtain the recombinant virus transfer vector, which was named pUCTKA-GFP-B. The homologous arms in the transfer vector are sequences on both sides of TK, so the recombinant virus obtained after recombination is a virus with TK gene deletion. Figure 4 It is a schematic diagram of the deletion position of the TK gene and the position of the TKA and TKB homology arms on the genome.

[0124] 1.1.1. Amplification and cloning of homologous recombination arm

[0125] 1.1.1.1 Primer design and template preparation

[0126] A...

Embodiment 3

[0212] Example 3 Preparation of PRV HN1201TK- / gE- / gI- / RR-deleted strain

[0213] 3.1 Construction of GFP intermediate transfer vector required for PRV HN1201 deletion RR

[0214] Prepare PRV HN1201TK with embodiment 2 - / gE - / gI - strain. According to the sequence of the fourth gene RR gene to be deleted, design homology arms at its two ends, namely RRA and RRB. RRA and RRB were cloned into pUC19 vector and named pUCRRAB. Then the GFP gene was cloned into pUCRRAB to obtain the recombinant virus transfer vector, named pUCRRA-GFP-B. The homologous arms in the transfer vector are the sequences on both sides of RR, so the recombinant virus obtained after recombination is the deletion of RR gene. The schematic diagram of recombinant transfer vector construction is shown in figure 1 , figure 2 is the location of the RRA and RRB homology arms on the genome.

[0215] 3.1.1 Amplification and cloning of homologous recombination arm

[0216] 3.1.1.1 Primer design and template...

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Abstract

The invention provides a porcine pseudorabies virus weak strain which is a porcine pseudorabies virus variant strain which can not express RR (response regulator) proteins. The invention further provides a vaccine composition containing a porcine pseudorabies virus weak strain antigen and a preparation method and application thereof. Through a live vaccine immunogenicity test and a pathogenic immunogenicity test, the porcine pseudorabies live vaccine is proved to have great protective efficacy, have basically no clinical symptoms and show great immune protection.

Description

technical field [0001] The invention relates to a porcine pseudorabies virus gene deletion strain, a prepared vaccine composition thereof, a preparation method and application thereof, and belongs to the field of animal virology. Background technique [0002] Pseudorabies, also known as Aujeszky's disease, is a disease of pigs, cattle, sheep and other domestic animals, poultry and wild animals caused by Suid herpesvirus 1 strain in the α subfamily of the Herpesviridae (Herpesviridae) family. An acute infectious disease characterized by fever, severe itching (except pigs) and encephalomyelitis. Pseudorabies in pigs is widespread in our country and is seriously harmful. It is one of the main diseases restricting the production of large-scale pig farms. It can cause abortion in pregnant sows, stillbirth or mummified fetuses and piglets with neurological symptoms, paralysis, and high mortality. PRV has strong pantropicity, neurotropicity, and latent infection characteristics. ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N7/00A61K39/245A61P31/22C12R1/93
Inventor 张许科孙进忠谭菲菲肖燕田克恭
Owner PULIKE BIOLOGICAL ENG INC
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