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1、METHODS25,402±408(2001)doi:10.1006/meth.2001.1262,availableonlineathttp://www.idealibrary.comonAnalysisofRelativeGeneExpressionDataUsingReal-2DDCTimeQuantitativePCRandthe2TMethodKennethJ.Livak*andThomasD.Schmittgen²,1*AppliedBiosystems,FosterCity,Califo
2、rnia94404;and²DepartmentofPharmaceuticalSciences,CollegeofPharmacy,WashingtonStateUniversity,Pullman,Washington99164-6534ofthetargetgenerelativetosomereferencegroupThetwomostcommonlyusedmethodstoanalyzedatafromsuchasanuntreatedcontrolorasampleattimezero
3、real-time,quantitativePCRexperimentsareabsolutequantifica-tionandrelativequantification.Absolutequantificationdeter-inatime-coursestudy.minestheinputcopynumber,usuallybyrelatingthePCRsignalAbsolutequantificationshouldbeperformedinsitu-toastandardcurve.R
4、elativequantificationrelatesthePCRsignalationswhereitisnecessarytodeterminetheabsoluteofthetargettranscriptinatreatmentgrouptothatofanothertranscriptcopynumber.Absolutequantificationhassamplesuchasanuntreatedcontrol.The22DDCTmethodisabeencombinedwithrea
5、l-timePCRandnumerousre-convenientwaytoanalyzetherelativechangesingeneexpressionportshaveappearedintheliterature(6±9)includingfromreal-timequantitativePCRexperiments.Thepurposeofthistwoarticlesinthisissue(10,11).Insomesituations,reportistopresentthederiv
6、ation,assumptions,andapplicationsofthe22DDCTmethod.Inaddition,wepresentthederivationanditmaybeunnecessarytodeterminetheabsolutetran-applicationsoftwovariationsofthe22DDCTmethodthatmaybescriptcopynumberandreportingtherelativechangeusefulintheanalysisofre
7、al-time,quantitativePCRdata.q2001ingeneexpressionwillsuffice.Forexample,statingElsevierScience(USA)thatagiventreatmentincreasedtheexpressionofKeyWords:reversetranscriptionpolymerasechainreaction;genexby2.5-foldmaybemorerelevantthanstatingquantitativepol
8、ymerasechainreaction;relativequantification;thatthetreatmentincreasedtheexpressionofgenexreal-timepolymerasechainreaction;TaqMan.from1000copiesto2500copiespercell.Quantifyingtherelativechangesingeneexpressionusingreal-timePCRrequ
