Construction And Preliminary Immunogenicity Assess Of Infectious Bronchitis Virus S1 Gene Suicidal DNA Vaccine Plasmid

In this reseach, the plamid pIBV—S1 (DQ288927)constructed by our lab was used toconstruct suicide DNA vaccine. The restrict site BamHⅠwas added by PCR. Then the S1gene of IBV was cloned and inserted into pSCA1. The recombinant plasmid pSCA1-S1was confirmed correct by PCR,restriction digestion and sequence analysis.Thelipospme-DNA IB DNA vaccines has been made at the ratio of 20/1.Recombinant plasmid pSCA1-S1,conventional DNA vaccine pcDNA3.1,emptyvector pSCA1 was transfected into Vero cells.48 hours later, expression of S1 in vitro wasverified by indirect immunofluorescence analysis. Recombinant plasmid pSCA1-S1,conventional DNA vaccine pcDNA3.1 can both be observed by green fluorescence, whilethe empty vector pSCA1 can not. The result confirm that the recombinant plasmidpSCA1-S1 can correttly express the S1 protein.7-day-old healthy chicken were injected intramuscularly with pSCA1-S1,conventional DNA vaccine pcDNA3.1,empty vector pSCA1,inactivated vaccine,PBScontral comparison. Picked the blood and separated the blood serum regularly afterimmunization continuously five weeks. Indirect ELISA assays were employed to evaluatehumoral immunity. The result showed that IBV-specific antibodies were induced at 7d afterimmunization. The data was maximum at the second week after immunization, thendeclined. The data between pSCA1-Sland conventional DNA vaccine pcDNA3.1 wassimilar. A significant difference was observed (P＜0.05), contrasting pSCA1-S1 with emptyvector. Fluorescence Activated Cell Sorter (FACS) were employed to evaluate cellimmunity. At 14d and 28d, the percent of CD3~+ was significant different by conventionalDNA vaccine pcDNA3.1(P＜0.05); at 21d,35 d, CD4~+ was significant different byconventional DNA vaccine pcDNA3.1(P＜0.05); CD8~+ was significant different byconventional DNA vaccine pcDNA3.1(P＜0.05 ) at 35d. The result shows that pSCA1-S1can induce a high level of cell immunity. Furthermore, the protected rate of pSCA1-S1, pcDNA3.1 was 60% and 50%. The result appraisal IBV suicide DNA vaccineimmunogenicity.This study constructed the infectious bronchitis virus S1 gene suicidal DNA vaccineplasmid, and assessed the preliminary immunogenicity. There is no report about that. Thisstudy should encourage further work towards the development of a new kind of DNAvaccine against IBV.