stepone-plus定量PCR操作手册

AppliedBiosystemsStepOne™andStepOnePlus™Real-TimePCRSystemsRelativeStandardCurveandComparativeCTExperimentsGettingStartedGuideAppliedBiosystemsStepOne™andStepOnePlus™Real-TimePCRSystemsRelativeStandardCurveandComparativeCTExperimentsGetStartedDesigntheRelativeStandardCurveExperimentPreparetheRelativeStandardCurveReactionsRuntheRelativeStandardCurveExperimentAnalyzetheRelativeStandardCurveExperimentDesigntheComparativeCTExperimentPreparetheComparativeCTReactionsRuntheComparativeCTExperimentAnalyzetheComparativeCTExperimentDesignandAnalyzeaStudyGettingStartedGuide©Copyright2008,2010AppliedBiosystems.Allrightsreserved.Informationinthisdocumentissubjecttochangewithoutnotice.AppliedBiosystemsassumesnoresponsibilityforanyerrorsthatmayappearinthisdocument.APPLIEDBIOSYSTEMSDISCLAIMSALLWARRANTIESWITHRESPECTTOTHISDOCUMENT,EXPRESSEDORIMPLIED,INCLUDINGBUTNOTLIMITEDTOTHOSEOFMERCHANTABILITYORFITNESSFORAPARTICULARPURPOSE.INNOEVENTSHALLAPPLIEDBIOSYSTEMSBELIABLE,WHETHERINCONTRACT,TORT,WARRANTY,ORUNDERANYSTATUTEORONANYOTHERBASISFORSPECIAL,INCIDENTAL,INDIRECT,PUNITIVE,MULTIPLEORCONSEQUENTIALDAMAGESINCONNECTIONWITHORARISINGFROMTHISDOCUMENT,INCLUDINGBUTNOTLIMITEDTOTHEUSETHEREOF.ForResearchUseOnly.Notforuseindiagnosticprocedures.NOTICETOPURCHASER:LabelLicenseTheStepOne™andStepOnePlus™Real-TimePCRSystemsarecoveredbyUSpatentsandcorrespondingclaimsintheirnon-UScounterparts,ownedbyAppliedBiosystemsInc.Norightisconveyedexpressly,byimplication,orbyestoppelunderanyotherpatentclaim,suchasclaimstoapparatus,reagents,kits,ormethodssuchas5′nucleasemethods.FurtherinformationonpurchasinglicensesmaybeobtainedbycontactingtheDirectorofLicensing,AppliedBiosystems,850LincolnCentreDrive,FosterCity,California94404,USA.TRADEMARKS:AppliedBiosystems,AB(Design),MicroAmp,PrimerExpress,andVICareregisteredtrademarks,andFAM,JOE,ROX,StepOne,StepOnePlus,TAMRA,andVeriFlexaretrademarksofAppliedBiosystemsInc.oritssubsidiariesintheU.S.and/orcertainothercountries.AmpErase,AmpliTaqGold,andTaqManareregisteredtrademarksofRocheMolecularSystems,Inc.Ambion,RNAlater,RNAqueous,RNaseZap,andSilencerareregisteredtrademarks,andDNAZap,Pre-miR,RiboPure,andTURBODNA-freearetrademarksofAmbion,Inc.SYBRisaregisteredtrademarkofMolecularProbes,Inc.MacintoshisaregisteredtrademarkofAppleComputer,Inc.MicrosoftandWindowsareregisteredtrademarksofMicrosoftCorporation.Allothertrademarksarethesolepropertyoftheirrespectiveowners.PartNumber4376785Rev.F06/2010ContentsiiiAppliedBiosystemsStepOne™andStepOnePlus™Real-TimePCRSystemsGettingStartedGuideforRelativeStandardCurveandComparativeCTExperimentsPreface.........................................viiHowtoUseThisGuide................................................viiHowtoObtainMoreInformation.........................................ixHowtoObtainSupport...............................................xiiiSafetyConventionsUsedinThisDocument...............................xivSymbolsonInstruments...............................................xvSafetyLabelsonInstruments...........................................xviiGeneralInstrumentSafety.............................................xviiiChemicalSafety.....................................................xixChemicalWasteSafety................................................xxElectricalSafety.....................................................xxiLEDSafety.........................................................xxiiBiologicalHazardSafety..............................................xxiiWorkstationSafety..................................................xxiiiSafetyandElectromagneticCompatibility(EMC)Standards..................xxiiiChapter1GetStarted...................................1AbouttheStepOne™andStepOnePlus™Systems............................2SupportedConsumables................................................4AboutRelativeStandardCurveandComparativeCTExperiments................6HowtoUseThisGuide................................................11AbouttheExampleExperiments.........................................12ExampleExperimentWorkflow..........................................18Chapter2DesigntheRelativeStandardCurveExperiment....21ChapterOverview.....................................................22CreateaNewExperiment..............................................23DefinetheExperimentProperties........................................27DefinetheMethodsandMaterials........................................30SetUptheTargets....................................................32SetUptheStandards..................................................35SetUptheSamples...................................................37SetUptheRelativeQuantitationSettings..................................39SetUptheRunMethod................................................41ReviewtheReactionSetup.............................................43AppliedBiosystemsStepOne™andStepOnePlus™Real-TimePCRSystemsGettingStartedGuideforRelativeStandardCurveandComparativeCTExperimentsivOrderMaterialsfortheExperiment..................................