Cell-mediated Immunity By S,N,E And M DNA Vaccine Of SARS-CoV

SARS(Severe Acute Respiratory Syndrome)is a contagious disease that has high infectivityand mortality as well as heavy damage on public health.The World Health Organization (WHO)announced in April 2003 that A novel coronavirus has been identified as the main causative agentof SARS. This virus is named SARS-Associated coronavirus (SARS-CoV). SARS-CoV is a newcoronavirus which entirely differs from other coronavirus. It has a high homogeneity withcoronaviruscomingfromanimalsandstrongvirulencewhichresultsinrespirationdisfunction,evendeath.Atpresent,thereisn'tspecifically effectivetherapy to cureviralinfection.Thebestmethod tocontrolthiskindofdiseaseisprevention.Amongthemethods, developingvaccineisakeystep.DNAvaccine with incomparable advantage is the third generation vaccine which follows pathogenyvaccine and subunit vaccine. Thus, this study choose the subject of SARS-CoV DNA vaccine andSpike glycoprotein, Nucleocapsid protein,Envelope protein and membrane protein gene ofSARS-CoV as candidate vaccine gene. We constructed recombinant DNA vaccine containingSARS-CoV structure gene and observed the production of SARS-CoV specific immune response inimmunizedmice,soastoinvestigatethefeasibilityofSARSvaccine.Thisstudyincludestwoparts.Inthefirstpart,weacquiredstructuregenesequenceofSARS-CoV(BJ01)fromGenBank.Accordingtothissequence,theprimersofstructuregenearedesignedbysoftwareprimer6.0. PCR amplified SARS-CoV structure gene at appropriate conditions. The amplified DNA wasdigested by restriction endonuclease,and ligated into pXAV1 vector. The recombinant plasmidincluding SARS-CoV structure gene was named pVAX/S,pVAX/N,pVAX/E,and pVAX/M. Therecombinant plasmid was identified by restriction endonuclease digestion and sequence analysis.ExpressionofstructureproteinwasdetectedbyIndirectImmunofluorescentMethod.In the second part,the recombinant plasmid was used to immunize the BALB/c miceintramuscularly.Mice were divided into three groups randomly:pVAX1 control plasmidgroup,testedgroups.After2,4,6,8,10weeks.TheCytokineofTNF,IFN-??,IL-5,IL-4,Il-2intheserumofmiceweredetectedwithFCM.The results showed that the recombinant plasmid DNA vaccine can induce specific cell-immunity and hormal immunity in mice.This might be benefit for further research on the safe andefficientSARSDNAvaccine.