Study On SE, S. Typhi From Ducks With Random Amplified Polymorphic DNA(RAPD), Cloning And Prokaryotic Expression Of OMP Gene From SE

This study concentrates on:1.Study on SE,S.typhi from Ducks with Random Amplified Polymorphic DNA(RAPD)The random amplified polymorphic DNA(RAPD),was used in this paper to study the genetic diversity among Salmonella enteritidis and S.typhimurium.The results showed that:(1)The arbitrary primers that can used by cluster analysising are A05,C14 and C17 in these arbitrary primers.(2)The length of all amplification products are between 0.2—0.3kb.(3)Cluster analysis are conducted foramplification products of three primers,and the results accorded with traditional classification.(4)Basically,it can cluster the isolates from different areas,but GH1,GH2,GH3 cluster with XJ firstly at similarity coefficient 0.9246,then cluster with GH4,5,6,7,8,9,10,11 at similarity coefficient 0.91382.Cloning and prokaryotic expression of OMP gene from SE Two pair primers were designed according to nucleotide sequence of SE which was submitted to GenBank with accession number of AF102556.The OMP gene was obtained by PCR with the SE isolated from ducks as template and cloned into pMD18-T vector.It was identified by PCR,restriction endonuclease analysis, DNA sequencing and then subcloned into EcoRⅠ/XhoⅠsite of prokaryotic expression vector pET-32a(+)and recombinant expression plasmid pET-32a-OMP was constructed successfully.The plasmid was transformed into Eschercha coli BL21(DE3)and 55kD OMP recombinant protein was expressed with IPTG inducer.The expression product were mainly non-soluble inclusion body.The recombinant protein was purified.The results of immunoblotting test showed that the recombinant protein had good immnuogenicity with using anti-SE rabbit sera.