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Brucellosis nucleic acid vaccine

A technology of brucella and nucleic acid vaccines, applied in the field of brucella nucleic acid vaccines, to achieve the effects of improving protection efficiency, ensuring yield and quality, and being easy to produce

Inactive Publication Date: 2009-10-14
PEKING UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, none of the vaccines described above are more protective than the currently used S19 or H38 vaccines

Method used

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  • Brucellosis nucleic acid vaccine
  • Brucellosis nucleic acid vaccine
  • Brucellosis nucleic acid vaccine

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033] Embodiment 1, the preparation of brucella vaccine

[0034] 1. Construction of eukaryotic expression vector:

[0035] The BCSP31, sodC and L7 / L12 genes of Brucella were amplified by PCR and cloned into the eukaryotic expression vector pJW4303 for expression in eukaryotic cells. GenBank accession numbers are: BCSP31: M20404; sodC: NC_007624; L7 / L12: L19101. The genomic DNA of Brucella abortus 2308 (purchased from China National Institute for the Control of Pharmaceutical and Biological Products) extracted by conventional methods was used as a template, and the BCSP31, sodC and L7 / L12 genes were amplified by PCR and cloned into the tissue on the pJW4303 vector Downstream of the plasminogen activator (TPA) signal sequence, a fusion protein is formed. The primer sequence used to amplify BCSP31 is as follows: 5'-GCTAGCCATATGAAATTCGGAAGCAAAATCCGTC-3' and 5'-CGGGATCCTTATTTCAGCACGCCCGCTTCC-3'; the primer sequence used to amplify sodC is as follows: 5'-GCTAGCCATATGAAAAAACGGCTT-...

Embodiment 2

[0037] Embodiment 2, identification of Brucella nucleic acid vaccine expressed in vivo

[0038] 1. RT-PCR identification of the expression of BCSP31, sodC and L7 / L12 genes

[0039] 20 C57BL / 6 mice were equally divided into two groups, and were treated as follows: the first group was the non-immune group (control), and each mouse was injected intramuscularly with 150 μl saline containing 150 μg pJW4303; the second group was treated with Brucella For the nucleic acid vaccine group, 50 μg of each of the plasmids pJBCSP31, pJsodC and pJL7 / L12 prepared in Example 1 were dissolved in 150 μl of normal saline and mixed evenly to obtain the Brucella nucleic acid vaccine. The brucella nucleic acid vaccine was immunized by intramuscular injection, and the immunization dose was 150 μl / bird. Four weeks after the completion of the immunization, spleens of 10 mice were taken from each group, and their total RNA was extracted using Trizol reagent (Gibco, USA). Then it was reverse transcribe...

Embodiment 3

[0045] Embodiment 3, the immune effect experiment of brucella nucleic acid vaccine:

[0046] 60 C57BL / 6 mice were equally divided into 4 groups, and were processed as follows: the first group was the Brucella nucleic acid vaccine group, each 50 μg of plasmids pJBCSP31, pJsodC and pJL7 / L12 prepared in Example 1 were dissolved together. In 150 μl of physiological saline, mix well to obtain the Brucella nucleic acid vaccine. The mice were immunized by intramuscular injection three times, the immunization dose was 150 μl / mouse / time, and the time points were the 0th week, the 3rd week, and the 6th week. The second group is the S19 vaccine group, 1×10 9 CFU S19 freeze-dried protein (purchased from China National Institute for the Control of Pharmaceutical and Biological Products, No. 040611, approval number: Xinxin Yaozi (2001) 287710) was dissolved in 150 μl of normal saline to obtain the S19 vaccine; the mice were immunized by intramuscular injection three times, and the immune ...

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Abstract

The invention discloses a Brucella nucleic acid vaccine. This brucella nucleic acid vaccine, its active ingredient is following any one mixture: 1) BCSP31, sodC and L7 / L12 gene are respectively cloned in the eukaryotic cell expression vector, obtain the recombinant expression vector containing BCSP31 gene, A recombinant expression vector containing the sodC gene and a recombinant expression vector containing the L7 / L12 gene, a mixture obtained by mixing the at least two recombinant expression vectors; 2) combining BCSP31, sodC and L7 / L12 with other nucleic acids After sequence fusion or partial sequence deletion or partial nucleotide mutation, the nucleotide sequences that can encode proteins with the same activity as BCSP31, sodC and L7 / L12 are respectively cloned into eukaryotic cell expression vectors, and the obtained at least two The mixture obtained by mixing the recombinant expression vectors. The brucella nucleic acid vaccine of the invention can be used for the prevention of human and animal brucellosis.

Description

technical field [0001] The invention relates to a brucella nucleic acid vaccine. Background technique [0002] Brucellosis is a zoonotic chronic wasting infectious disease worldwide, caused by a Gram-negative facultative intracellular bacterium named Brucella, in humans, cattle, sheep, Rodents, pigs and other mammals are widely spread. In species important to animal husbandry, the disease can greatly lead to the loss of animal reproductive ability; for humans, there will be clinical symptoms such as intermittent fever, so it is also called wave fever. Vaccine immunization is essential to combat Brucellosis. S19 live attenuated bacterial vaccine is the earliest and most widely used vaccine in cattle herds. Although the vaccine conferred some protection, its persistent serological response made it impossible to distinguish immunized from infected cattle. In addition, the vaccine is pathogenic to humans and can cause abortion in pregnant cattle. Similar vaccines such as Re...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61K48/00A61K39/10A61P31/04C12N15/79
Inventor 蔡宏朱玉贤
Owner PEKING UNIV
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