Rapid and High-Throughput pan-Orthopoxvirus Detection and Identification using PCR and Mass Spectrometry 英文参考文献.docVIP

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Rapid and High-Throughput pan-Orthopoxvirus Detection and Identification using PCR and Mass Spectrometry 英文参考文献.doc

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Rapid and High-Throughput pan-Orthopoxvirus Detection and Identification using PCR and Mass Spectrometry 英文参考文献

RapidandHigh-Throughputpan-Orthopoxvirus DetectionandIdentificationusingPCRandMass Spectrometry MarkW.Eshoo1*,ChrisA.Whitehouse2,AysegulNalca2,ScottZoll1,JosephA.Ecker1,ThomasA.Hall1, Thuy-TrangD.Pennella1,DavidD.Duncan1,AnjaliDesai3,EmilyK.Moradi3,KarlRudnick3,BrianLibby1, RaymondRanken1,RangarajanSampath1,StevenA.Hofstadler1,DavidJ.Ecker1,LawrenceB.Blyn1 1IbisBiosciences,Carlsbad,California,UnitedStatesofAmerica,2UnitedStatesArmyMedicalResearchInstituteofInfectiousDiseases,FortDetrick,Maryland,United StatesofAmerica,3ScienceApplicationsInternationalCorporation(SAIC),SanDiego,California,UnitedStatesofAmerica Abstract ThegenusOrthopoxviruscontainsseveralspeciesofrelatedviruses,includingthecausativeagentofsmallpox(Variolavirus). Inadditiontosmallpox,severalothermembersofthegenusarecapableofcausinghumaninfection,includingmonkeypox, cowpox, and other zoonotic rodent-borne poxviruses. Therefore, a single assay that can accurately identify all orthopoxviruses could provide a valuable tool for rapid broad orthopovirus identification. We have developed a pan- OrthopoxvirusassayforidentificationofallmembersofthegenusbasedonfourPCRreactionstargetingOrthopoxvirusDNA and RNA helicase and polymerase genes. The amplicons are detected using electrospray ionization-mass spectrometry (PCR/ESI-MS) on the Ibis T5000 system. We demonstrate that the assay can detect and identify a diverse collection of orthopoxviruses,providesub-speciesinformationandcharacterizevirusesfromthebloodofrabbitpoxinfectedrabbits.The assayissensitiveatthestochasticlimitofPCRanddetectedvirusinbloodcontainingapproximatelysixplaque-forming unitspermilliliterfromarabbitpoxvirus-infectedrabbit. Citation:EshooMW,WhitehouseCA,NalcaA,ZollS,EckerJA,etal.(2009)RapidandHigh-Throughputpan-OrthopoxvirusDetectionandIdentificationusingPCR andMassSpectrometry.PLoSONE4(7):e6342.doi:10.1371/journal.pone.0006342 Editor:DingXiangLiu,InstituteofMolecularandCellBiology,Singapore ReceivedFebruary11,2009;AcceptedJune1,2009;Publ