河北豬繁殖與呼吸綜合征病毒地方株ORF5基因的原核表達.pdf

1、河北農業(yè)大學碩士學位論文河北豬繁殖與呼吸綜合征病毒地方株F5基因的原核表達姓名：劉濤申請學位級別：碩士專業(yè)：預防獸醫(yī)學指導教師：孫繼國20080607ExpressionoftheORF5geneofPorcineReproductiveandRespiratorySyndromeVirusinHeBeiCandidate：LuTaoSpecialt)r：PreventiveVeterinaryMedicineAdvisor：SunJi

2、guoAbstractPorcinereproductiveandrespiratorysyndromeisanewinfectiousdiseaseItcausesabortionsinSOWSandrespiratoryproblemsinpi酉etsItwascausedbyporcinereproducfiveandrespiratorysyndromevims(PRRSV)Sinceit’Sappearance，PRRShas

3、beencausingtremendouseconomiclossestotheswineindustrythroughouttheworldItisdi伍culttocontrolPRRSwithoutalleffectivevaccineItisevidentthatthecurrentvaccinesarenoteffectiveinprotectingagainstinfections、Ⅳimthegeneticallydive

4、rsefieldstrainsofP砌己SVTheporcinereproduetiveandrespiratorysyndromevirusisolatedfromtheclinicsuspeetedlungmaterial，namedHB一3(cz)Postinoculationmanytimes，theCPEinMare145cellappearedinroundinggatheringsheddingfeaturesstably

5、AccordingtothecompletenucleotidesequenceofPRRSVCHlainOenBankapairofprimersweredesignedtoamplifyORF5geneofliB3(cz)strain11lespecificORF5geneamplifiedbyRTPCRwasclonedintopMD—Tvemor功erecombinantplasmidWassequencedandcompare

6、dwimotherstrains耽ehydrophobicitytransmembranehelices，signalpeptideandlatentantigensiteofGP5proteinarehomologousofallHebeistrainsTheORF5genehave896％and891％homologies麗mVR2332(aNorthAmericanisolate)innucleotideandanlineacid

7、level，whereasithasonly637％and532％homologieswithLV(anEuropenisolate)；T11eHB一3(cz)strainbelongtoNorthAmericangenotype；ThephylogenetictreesrevealedthatthestrainsisolatedinrecentyearsareclosedwimHB—l(sh)strainThedORF5geneseg

8、mentdeletingNterminalhydrophobicsequenceWaSamplifiedfromORF5geneofHB一3(cz)strainbyPCR，thenthegenewasclonedintoprokaryoticexpressionvectorpGEX6P1TherecombinantfusionproteinsdGP5werehighlyexpressedinE—coliBL21intheformsofi

9、nclusionbodiesinducedbyIPTGWeSternblottingshowedthattherecombinantproteincouldreactwiththeporcinepolyclonalantibodyagainstPRRSVindicatingthattheproteincouldbeusedasantigenindiagnostieassayforthedetectionofantibodiesagain