含有Th细胞表位的合成PCV2 ORF2 Cap多肽抗原的表达及免疫原性分析

doi:10.11843/j.issn.0366-6964.2015.12.020

Abstract

Abstract:

To determine the role of Th cell epitopes，which were screened from PCV2 ORF1，and ORF3，in improving the PCV2 ORF2 Cap P1 immunogenicity when they were added to the sequence，and the feasibility of synthetic expression of PCV2 ORF2 Cap P1 as a vaccine antigen，bioinformatics and molecular biology methods were employed to screen the peptide sequences.A pan-host of T helper cell epitope peptides and three Th cell epitopes peptide sequences of PCV2-specific were identified.These 4 Th cells peptides sequences were combined with a B-cell epitope peptide sequence screened from PCV2 ORF2 Cap1 gene，and then codon was optimized and inserted the restriction sites and stop codons.After analyzing the codon adaptation index and the frequency distribution，predicted it can achieve efficient expression，further chemical synthesis or peptides were carried out.The synthetic polypeptide antigen P1 was connected to the pET-30a expression vector，and transformed into BL21(DE3) pLysS competent cells，the expression in the engineered bacteria BL21(DE3) pLysS-pET-30a-P1 was constructed.The P1 antigen was able to be expressed after IPTG inducting.The expression level was identified by SDS-PAGE，and its biological activity was identified by Western Blot，followed by mice and pigs immunization with P1.Antibody levels in mice and the peripheral blood lymphocyte proliferation in pigs were tested.The results showed that the codon adaptation index of PCV2 P1 sequence was 0.89，it can achieve to high levels of expression，and the peptide fragment is about 28.29 kDa.MTT results showed that P1 peptide antigen had good immunogenicity.In immunized pigs，the expression of IFN-γ and IL-4 can increase obviously，and were significantly different to the control group(P<0.05).These results indicated that P1 could induce a strong humoral and celluar immune response，and it will provide a theoretical basis on PCV2 P1 peptide antigen as a vaccine antigen for further studies.

CLC Number:

S852.4

CHEN Shan-zhen，ZHAO Yan，LI Zhong-sheng，LUO Jun，CHEN Ke-hong，WANG Gui-ping，LI Qi-chang. Expression and Immunogenicity Analysis of PCV2 Cap Recombinant Protein Antigen which Contains Helper T cell Epitope Peptides[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2015, 46(12): 2273-2281.

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Expression and Immunogenicity Analysis of PCV2 Cap Recombinant Protein Antigen which Contains Helper T cell Epitope Peptides

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