Porcine circovirus type 3 virus-like particle as well as preparation method and application thereof

Abstract

The invention provides a porcine circovirus type 3 virus-like particle as well as a preparation method and application thereof. According to the invention, an insect-baculovirus expression system is utilized to express a PCV3 Cap protein variant so as to prepare virus-like particles. According to the invention, double promoters of baculovirus are used to start Cap protein expression, on the basis of keeping the N terminal of the Cap protein rich in basic amino acid, the first to 22th amino acids are deleted, and a sequence for coding the basic protein P6.9 N terminal rich in basic amino acid of the baculovirus is inserted, so that the Cap protein is expressed in a fusion manner. Results show that the expressed fusion protein not only can form the virus-like particles, but also can be expressed at a relatively high level, and a basis is provided for further development of PCV3 vaccines and realization of industrial production of the PCV3 virus-like particles.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a porcine circovirus type 3 virus-like particle and a preparation method and application thereof. Background technique [0002] Porcine circovirus (Porcine circovirus, PCV) belongs to Circoviridae (Circoviridae) genus Circovirus (Circovirus), contains a single strand of circular DNA, is one of the smallest animal DNA viruses, first in 1974 in pig kidney cells line (PK-15 cell line). At present, four genotypes of PCV have been found, namely PCV1, PCV2, PCV3, and PCV4. PCV1 is non-pathogenic, and PCV4 is a new genotype discovered in 2019, and its pathogenicity needs further study. PCV2 can infect monocytes, macrophages, etc., causing immune cell damage in pigs, leading to severe immunosuppression in infected pigs, resulting in postweaning multisystemic wasting syndrome (PMWS) and dermatitis and nephrotic syndrome (PDNS). , proliferative necrotizing interstitial pneumonia...

AI Technical Summary

Problems solved by technology

At present, although the full-length and N-terminal 32 amino acid truncated Cap proteins are expressed in baculovirus and can form VLP (virus-like particle), the amino acids 1-32 of the N-terminus of the Cap protein are rich in basic Amino acid, which can mediate the nuclear transport of Cap protein, the expression of the complete Cap protein may affect its expression in the expression system, but the deletion of the entire segment may affect the characteristics of the surface of the virus-like particle and the characteristics of the virus-like particle Stability, thus affecting its immunogenicity

Initial research found that the expression level of the complete PCV3 Cap protein and the Cap protein lacking 32 amino acids at the N-terminal is extremely low in insect cells, which cannot meet the needs of industrialization

In addition, in the above scheme using the baculovirus expression system, only the codon-unoptimized Cap gene was inserted into the expression vector, and no other elements were added or the vector was modified to further increase the expression of the protein.

[0005] In summary, the research and development of PCV3 Cap virus-like particles is of great significance, but in the current expression of Cap protein, the process of prokaryotic expression system is relatively complicated, the protein has no post-translational modification, and the immunogenicity of virus-like particles obtained is poorer than that of eukaryotic expression. , while the yeast expression system has low secretion expression, and the insect-baculovirus expression system still needs to be further optimized and improved

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