Preparation And Immunogenicity Of The Asia Type I Foot-and-mouth Disease Virus Genetically Engineered Peptide Vaccine Analysis

Chapter 1 Analysis of the VP1 nucleotide sequence of foot-and-mouth disease virus serotype Asiaâ… AKT strainFor constructing a novel recombinant peptide vaccine,we isolated RNA of FMDV serotype Asiaâ… AKT strain and reverse-transcripted into cDNA.The PCR product was then cloned to pMD19-T vector.The nucleotide sequence of VP1 was determined by dideoxy-mediated chain termination method.Compared the differences and homologies of VP1 nucleotide sequence and amino acids sequence between AKT strain and the other strain of FMDV Asiaâ… serotype such as India,YNBS,and jiangsu strain using DNAStar software.The analysis results show that the homologies of nucleotide sequences between AKT strain and India,YNBS,and jiangsu strains are 92.4%,83.3%,84.5%,the relevant homologies of amino acids sequences are 94.8%, 91.0%,89.6%.Analysising the VP1 gene sequences of FMDV Asiaâ… serotype provides important data for the research of studing the VP1 gene mutation and designing an effective vaccine against foot-and-mouth disease virus serotype Asiaâ… . Chapter 2 The recombinant peptide vaccines against foot-and-mouth disease virus serotype Asiaâ… and their efficacy in guinea pigsWe get the nucleotide sequences of 133-163 amino acids of VP1 protein of foot-and-mouth disease virus serotype Asiaâ… from chapter 1,and the 1-33 amino acids of VP2 protein from GenBank,then amplified the DNA fragments and ligated them toβ-galactosidase gene 3' terminal respectively constructing two recombinant expression plasmids named pLacZ'-VP1 and pLacZ'-VP2.The two epitopes were ligated into a tandem sequence as VP1(133-163)-VP2(1-33)-VP1(133-163) and chemically synthesized,then ligated the tandem sequence toβ-galactosidase gene to construct an expression plasmid named pT1.In the same way we ligated the tandem sequence to the bovine IgG heavy chain constant region to construct another expression plasmid pT2.The efficacy of the vaccines were investigated by neutralization antibody assay,lympho-proliferation assay and viral challenge assay.The results showed that both pT1 and pT2 fusion proteins can significantly stimulate cellular and humoral response.Our result of viral challenge showed that pT1 and pT2 could protect guinea pigs with 100%protection rate against 100 ID₅₀ viral challenge with FMDV serotype Asiaâ… by the twice vaccination method,but then only 70%by one time vaccination with pT2.