Preparation method of recombinant lactobacillus plantarum expressing porcine circovirus type-3 Cap gene

Abstract

The invention relates to the technical field of biological medicine engineering and discloses a preparation method of recombinant lactobacillus plantarum expressing a porcine circovirus type-3 (PCV3)Cap gene. The method comprises the following step of S1, design and synthesis of primers, wherein the primers LpS-F and LpS-R for amplifying the sequence of the PCV3 Cap gene are designed through Primer 5.0 software according to a PCV3 gene sequence (login number: AYM55798.1) in GenBank, and the fragment length is 789bp. According to the preparation method of recombinant lactobacillus plantarum expressing the porcine circovirus type-3 Cap gene, lactobacillus plantarum is selected to express the PCV3 Cap gene which contains the signal peptide 1320 and is optimized by DCpep, lactococcus NZ3900 is selected as a cloning intermediate host, and a high content of recombinant plasmids are obtained; then, lactobacillus plantarum Lp12 is used for expressing the PCV3 Cap protein, an effective preparation and identification method is established, a detection means is optimized, and a theoretical foundation is laid for developing type-3 porcine circovirus candidate vaccines taken orally. By means of the preparation method, recombinant lactobacillus plantarum for expressing the PCV3 Cap protein is created, and the feasibility of expressing heterologous proteins through lactobacillus plantarum isenriched.

Description

technical field [0001] The invention relates to the technical field of biomedical engineering, in particular to a preparation method of a recombinant plant lactic acid bacteria expressing porcine circovirus type 3 Cap gene. Background technique [0002] Porcine circovirus is a single-stranded circular DNA virus of the genus Circovirus in the family Circoviridae; the known genotypes of the virus are PCV1, PCV2 and PCV3. PCV1 is a cellular contaminant and is not pathogenic. PCV2 is the main causative agent of related diseases such as post-weaning multisystemic wasting syndrome, porcine dermatitis and nephrotic syndrome, and reproductive disorders. This PCV2-related disease is named as porcine circovirus-related disease. In 2016, American researchers used metagenomic sequencing to discover porcine circovirus type 3, which clinically manifested as PDNS-like, reproductive disorders, and multisystem inflammation. At present, PCV3 has been widely spread in the world. Zheng et al....

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Problems solved by technology

[0004] At present, the genetically engineered subunit vaccines against PCV2 on the market all use Cap protein as the target protein. Ni Nengneng et al. expressed PCV2Cap protein by Lactobacillus plantarum, but there is no report on the expression of PCV3 Cap protein by Lactobacillus plantarum. According to genome sequence analysis, it was found that PCV3 The similarity with PCV2Cap protein is only 37%, and it is closer to bats or dogs in terms of genetic relationship. Whether the vaccine against PCV2Cap protein can produce cross-protection against PCV3 is worth considering, so the vaccine against PCV3 still needs to be further invented

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