Bacterin for pleuropneumonia actinobacillus serotype 1 double-gene deletion mutant without resistance marker

A technology of porcine pleuropneumonia and actinobacillus, applied in the direction of antibacterial drugs, bacteria, bacterial antigen components, etc., can solve the problems of not meeting the biological safety requirements, not being able to be used as vaccine strains, etc., and achieve the protection of homologous and heterologous serotypes Bacteria attack, broad market application prospects, strong virulence effect

Inactive Publication Date: 2007-09-19
HUAZHONG AGRI UNIV
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Problems solved by technology

For example, there is randomness in chemical or transposon-mediated mutation methods, which are only suitable for the screening of mutants with obvious phenotypic changes, and mutant strains that do not cause obvious phenotypic changes cannot be obtained by these

Method used

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  • Bacterin for pleuropneumonia actinobacillus serotype 1 double-gene deletion mutant without resistance marker
  • Bacterin for pleuropneumonia actinobacillus serotype 1 double-gene deletion mutant without resistance marker
  • Bacterin for pleuropneumonia actinobacillus serotype 1 double-gene deletion mutant without resistance marker

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Embodiment 1

[0042] 1. Primer design (for gene cloning and molecular detection)

[0043] According to the reported sequence of APP-1 strain (refer to the gene sequence of GenBank accession number X68595), four pairs of primers were designed to amplify the upstream arm and downstream arm of the toxin apxI activating gene apxIC, respectively. The size of the amplified fragments were 800bp and 1700bp, and the upstream arm Both ends are designed with EcoRI and HindIII restriction sites, and both ends of the downstream arm are designed with HindIII and XhoI restriction sites; the upstream and downstream arms of the activation gene apxIIC of apxII, the amplified fragment size is 1600bp and 1800bp, respectively, the upstream arm PstI and EcoRI restriction sites were designed at both ends, and KpnI and XhoI restriction sites were designed at both ends of the downstream arm. The above primers were all synthesized by Shanghai Bioengineering Company. The primer sequence is as follows:

[0044] pI-1: 5’-A...

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Abstract

The invention belongs to animal bacterium gene engineering field, relates to form stem of Actinobacillus pleuropneumoniae APP-1-mut01 digene deletion mutants without fastness label, preparing vaccine and application thereof. The invention gets a stem of Actinobacillus pleuropneumoniae APP-1-mut01 (storing number is CCTCC NO; M207005). The stem absents two main toxin gene activity factors apxIC and apxIIC of Actinobacillus pleuropneumoniae APP-1-mut01, produces toxin albumen ApxIA and ApxIIA without toxin, but these two toxin albumen also have immunity originality. The invention also discloses producing vaccine of Actinobacillus pleuropneumoniae APP-1-mut01 using the digene deletion mutants stem and application thereof. The digene deletion vaccine can irritate pig generate protective immunity reaction to resist isogeny and nonhomology serum wild strain of actinobacillus pleuropneumoniae to prevent infection thereof.

Description

Technical field [0001] The invention belongs to the technical field of animal bacterial genetic engineering, and specifically relates to the construction, vaccine preparation and application of a double gene deletion mutant strain of Actinobacillus pleuropneumoniae serotype 1 without resistance markers Background technique [0002] Porcine contagious pleuropneumonia (PCP) is an infectious respiratory disease of pigs caused by Actinobacillus pleuropneumoniae (APP), which has caused serious economic losses to the world pig industry. Since the discovery of the PCP epidemic in my country, the majority of veterinarians have conducted a series of studies on its etiology, epidemiology, diagnosis and control, and achieved certain achievements. However, the incidence of this disease in my country is still increasing year by year, and the positive rate of some pig farms has reached more than 70%. It has become one of the main infectious diseases in intensive pig far...

Claims

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Application Information

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IPC IPC(8): C12N1/21A61K39/02A61P31/04
Inventor 贝为成陈焕春林丽雯刘金林周锐金梅林何启盖方六荣吴斌肖少波郭爱珍曹胜波
Owner HUAZHONG AGRI UNIV
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