The Study On Plague Sub-unit And Oral Live Vector Vaccine

Y.pestis is the etiological agent of bubonic and pneumonic plague, diseases which have caused over 200 million human deaths in the past. Plague still occurs throughout the world today, though for reasons that are not fully understood pandemics of disease do not develop from these outbreaks. Antibiotic treatment of bubonic plague is usually effective, but pneumonic plague is difficult to treat and even with antibiotic therapy death often results. A killed whole cell plague vaccine has been used in the past, but recent studies in animals have shown that this vaccine offers poor protection against pneumonic disease. A live attenuated vaccine is also available. Whilst this vaccine is effective, it retains some virulence and in most countries it is not considered to be suitable for use in humans. A sub-unit vaccine based on the F1 and V antigens is highly effective against both bubonic and pneumonic plague, when tested in animal models of disease. This vaccine has been used to explore the utility of different intranasal and oral delivery systems, based on the Salmonella delivery of sub-units.DNA vaccine also is hopeful to plague prevention. We try here to develop improved sub-unit and live attenuated vector vaccines against plague.In this study,1.F1 gene and V gene fragment of Y pestis EV76 was amplified by polymerase chain reaction (PCR) and fusion cloned and inserted into pGEM-T vector, After sequence correction, the recombinant expression plasmid F1-V/11C was constructed by inserting the DNA fragment of Y pestis F1 gene and V gene into pET-11c and transformed into E coli BL21 cell. The positive clone was selected by PCR and enzymes digest and the expression product of F1-V gene induced by IPTG was detected by SDS-PAGE, while the expression protein immunogenicity was detected by Western-blot. Different purification strategies were used to purify F1-V/11C. First hydrophobic chromatograph was taken, second anion exchange chromatograph was used at last size exclusion chromatograph was adapted, and then alhydrogel-absorbed F1-V fusion protein has been prepared to intramuscularly inject Balb/c mice.2.F1-V gene were amplified by PCR and cloned into a prokaryotic expression plasmid asd-pTrc99A.The recombinant plasmid was identified and then used to transform the...