African swine fever virus chimeric protein, vaccine composition, preparation method and application thereof
A technology of African swine fever virus and vaccine composition, which is applied in the direction of biochemical equipment and methods, viruses, vaccines, etc., and can solve problems such as unsatisfactory vaccine immune effects
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Embodiment 1
[0057] Example 1 Expression of African swine fever virus p72 full-length and truncated proteins
[0058] The nucleotide sequence of the amino acid sequence of the p72 protein shown in SEQ ID NO.1 of the corresponding coding sequence table was synthesized by Suzhou Jinweizhi Biotechnology Co., Ltd., and the synthesized nucleotide was cloned into the pET28a vector, named as the recombinant plasmid pET28a-p72 .
[0059] Similarly, the African swine fever virus p72 domain I shown in the sequence table SEQ ID NO.2, the African swine fever virus p72 domain II shown in the sequence table SEQ ID NO.3, and the African swine fever virus shown in the sequence table SEQ ID NO.4 Pestivirus p72 domain III is connected by a flexible linker peptide, in which African swine fever virus p72 domain I is located at the N-terminus, domain III is located at the C-terminus, and domain II is located between domains I and III. Weizhi Biotechnology Co., Ltd. synthesized the corresponding coding nucleot...
Embodiment 2
[0063] Embodiment 2 The impact of African swine fever virus p72 truncated protein on immunogenicity
[0064] The p72 full-length protein and p72 truncated protein prepared in Example 1 were added to the adjuvant, and the process of adding was continuously stirred with an emulsifier at a speed of 800 rpm for 12 minutes, mixed, and stored at 4°C, that is, the p72 full-length protein or p72 truncated protein Vaccine compositions of p72 truncated proteins. The adjuvants suitable for the present invention may be adjuvants known to those skilled in the art. In the present invention, the adjuvant selected is a biphasic adjuvant (water-in-oil-in-water emulsion), for example, the adjuvant ISA206 (Seppic, France). The specific ratio of each component in the prepared vaccine is shown in Table 1.
[0065] Table 1 African swine fever virus p72 full-length and truncated protein vaccine composition composition ratio
[0066] components Vaccine 1 Vaccine 2 p72 protein (μ...
Embodiment 3
[0080] Embodiment 3 African swine fever virus chimeric protein backbone and antigenic protein selection
[0081] The inventor analyzed the structure of the African swine fever virus p72 protein and creatively truncated it, which still maintained the relative stability of the protein space structure, and its immunogenicity was not affected. Therefore, the p72 truncated protein was determined as the embedding protein of the present invention. The skeleton of the protein.
[0082] At the same time, after a large number of experimental studies, the inventors have screened the following sequence in combination with the antigenic epitope analysis of the African swine fever virus immunogenic protein. Fragments of at least one, to produce a higher immunogenic response:
[0083] The p54 protein fragment is selected from any one or more of SEQ ID NO.5, 6, 7, 8, 9, 10, 11, 12;
[0084] The p30 protein fragment is selected from any one or more of SEQ ID NO.13, 14, 15, 16, 17, 18, 19;
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