Oneoftheviewsavailableaftercompletionoftherunisanamplificationwindow.Thiswindowshowstheamountoffluorescenceobtainedineachamplificationcycleforeachreaction.Thethresholdcycle(Ct)isshownbythedarkerhorizontalline.第55页，共103页，星期日，2025年，2月5日AdvantageProvidesanaccuratemethodfordeterminationoflevelsofspecificDNAandRNAsequencesintissuesamples.BasedondetectionofafluorescentsignalproducedproportionallyduringamplificationofaPCRproduct.Turn-aroundtimefordataacquisitionandanalysisisshort,ResultsaremorereliablethanbytraditionalPCRmethods.SybrgreenIdetectionislessexpensiveandnosequencespecificprobeisrequired.Userswillalsoneedtopurchaseflat-top,opticalcapsor0.2mmthermalmicrosealfilmfortheirplates.Allotherplatesandcapswillnotworkwiththenewmachines.第56页，共103页，星期日，2025年，2月5日ApplicationsTheapplicationsforquantitativereal-timePCRareinnumerable（数不清的）.DetectionofgenomicorviralDNAintissuescanbeavaluablediagnostictool.GeneexpressioncanbemeasuredafterextractionoftotalRNAandpreparationofcDNAbyareversetranscription(RT)step.SetupandanalysisaresimpleandcanmoreeasilybeextendedtotheclinicalenvironmentthantraditionalPCRtechniques.Anallelicdiscriminationassaythatcandetectsingle-basenucleotidemutationsandpolymorphisms.第57页，共103页，星期日，2025年，2月5日Allelic(等位基因的)discriminationassayTheseassaysrequiretwoseparateprobesthatdifferonlybyonebasemismatch.Oneprobelabeledwith6-FAM(6-羧基荧光素)representsoneallele（等位基因）,andtheotherprobelabeledwiththefluorochromeVICrepresentstheotherallele.Amismatchleadstoalessefficientamplification.Fluorescencespectraarecollectedaftertherun,andusingmulticomponentanalysis,thesoftwareextractsthecontributionofeachcomponentdyetotheobservedspectrum.第58页，共103页，