Study On Expression Of Human Cytomegalovirus Triple Peptide And Its DNA Vaccine

To express the triple-peptide of gp52 pp150 and gB C-terminal peptide from human cytomegalovirus(HCMV) in Pichia pastoris, to construct its DNA vaccine and observe the immune responses in vaccined rabbit.The expression vector pPIC9K-6His-HCMV3p containing the interesting gene fragment was constructed and transformed into the Pichia pastoris. The recombinant triple-peptide was then expressed and purified by Ni²⁺-chelate affinity chromatography. In addition, the gene of the triple- peptide was cloned into the eukaryotic expression plasmid pVAX1 and the eukaryotic recombinant expression vector pVAX1-HCMV3p was obtained, then they were used to immunize rabbits via muscles. The triple-peptide specific IgG in serum was analyzed by indirect enzyme-linked immunosorbent assay (ELISA). Using indirect fluorescence assay(IFA) to detect HCMV-specific IgG The proliferation response of PBMCs was tested by MTT assay and the HCMV specific lymphocytes response by LDH release test.Both SDS-PAGE and Western-blot showed that the recombinant triple-pepide has been expressed in Pichia pastoris successfully. The recombinant triple-peptide reacted specifically with the sera from immunized rabbit. The DNA vaccine could induce specific triple-peptide antibodies and specific HCMV antibodies in rabbit. The specific proliferation activity and anti-virus responses of PBMCs from the rabbit immunized with DNA vaccine were observed.It is concluded that the recombinant HCMV triple- peptide could be used as a antigen to detect antibodies for HCMV infection; Specific cellular and humoral immune responses in rabbit can be induced with its DNA vaccine, which is useful for developing HCMV vaccine.