冠状病毒Coronavirusl论文-2016 [Springer Protocols Handbooks] Animal Coronaviruses __ A Multiplex Polymerase Chain Reaction for Differential Detec.pdf

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1、Chapter12AMultiplexPolymeraseChainReactionforDifferentialDetectionofTurkeyCoronavirusfromChickenInfectiousBronchitisVirusandBovineCoronavirusChienChangLoa,ChingChingWu,andTsangLongLinAbstractAmultiplexpolymerasechainreaction(PCR)methodfordifferentialdetectionofturkeycoronavirus(TCoV),infecti

2、ousbronchitisvirus(IBV),andbovinecoronavirus(BCoV)ispresentedinthischapter.Primersaredesignedfromtheconservedorvariableregionsofnucleocapsid(N)orspike(S)proteingenesofTCoV,IBV,andBCoVandusedinthesamePCRreaction.ReversetranscriptionfollowedbyPCRreactionisusedtoamplifyaportionofNorSgeneoftheco

3、rrespondingcoronaviruses.TwoPCRprod-ucts,a356-bpbandcorrespondingtoNgeneanda727-bpbandcorrespondingtoSgene,areobtainedforTCoV.Incontrast,onePCRproductof356bpcorrespondingtoafragmentofNgeneisobtainedforIBVstrainsandonePCRproductof568bpcorrespondingtoafragmentofSgeneisobtainedforBCoV.KeywordsM

4、ultiplexPCR,Differentialdetection,Coronavirus,Nucleocapsidgene,Spikegene1IntroductionTurkeycoronavirus(TCoV)contributedtosigniÞcanteconomiclossesandremainsasaseriousthreattotheturkeyproducers.Turkeycoronaviralenteritisinareaswithhighconcentrationsofturkeysonayear-roundbasisisnoteasilyelimina

5、tedandisencounteredfrequentlyinturkeypoults[1].AccurateandrapidmethodfordiagnosisofTCoVinfectionisthekeytoeffectivecontrolofthedisease.TurkeycoronavirusbelongstothefamilyCoronaviridae,whichisagroupofenveloped,positive-strandedRNAvirusesthatinfectawiderangeofmammalianandavianspecies.Themajors

6、tructuralproteinsofcoronavirusincludephosphorylatednucleocapsid(N)protein,peplomericspike(S)glycoprotein,andtransmembraneormembrane(M)glycoprotein.Spikeproteincontributestothedistinctivepeplomersontheviralsurfaceandcontainsneutralizingandgroup-speciÞcepitopes.Spikeproteinishighlyvariableamon

7、gLeyiWang(ed.),AnimalCoronaviruses,SpringerProtocolsHandbooks,DOI10.1007/978-1-4939-3414-0_12,©SpringerScience+BusinessMediaNewYork2016131132ChienChangLoaetal.differentcoronaviruseswhileMandNproteinsaremoreconservedamongcoronavirusesbetweendifferen