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1、MINIREVIEWAdvantagesandlimitationsofquantitativePCR(Q-PCR)-basedapproachesinmicrobialecologyCindyJ.Smith&A.MarkOsbornDepartmentofAnimalandPlantSciences,UniversityofSheffield,WesternBank,Sheffield,UKCorrespondence:A.MarkOsborn,AbstractDepartmentofAnimalandPlantSciences,QuantitativePCR(Q-PCRorreal-time
2、PCR)approachesarenowwidelyappliedUniversityofSheffield,WesternBank,SheffieldS102TN,UK.Tel.:144114222inmicrobialecologytoquantifytheabundanceandexpressionoftaxonomicand4626;fax:1441142220002;e-mail:functionalgenemarkerswithintheenvironment.Q-PCR-basedanalysescombinea.m.osborn@sheffield.ac.uktraditional
3、end-pointdetectionPCRwithfluorescentdetectiontechnologiestorecordtheaccumulationofampliconsinrealtimeduringeachcycleofthePCRReceived20June2008;revised10Octoberamplification.Bydetectionofampliconsduringtheearlyexponentialphaseofthe2008;accepted24October2008.PCR,thisenablesthequantificationofgene(ortran
4、script)numberswhentheseFirstpublishedonlineDecember2008.areproportionaltothestartingtemplateconcentration.WhenQ-PCRiscoupledwithaprecedingreversetranscriptionreaction,itcanbeusedtoquantifygeneDOI:10.1111/j.1574-6941.2008.00629.xexpression(RT-Q-PCR).Thisreviewfirstlyaddressesthetheoreticalandpractica
5、limplementationofQ-PCRandRT-Q-PCRprotocolsinmicrobialecology,high-Editor:MichaelWagnerlightingkeyexperimentalconsiderations.Secondly,wereviewtheapplicationsofKeywords(RT)-Q-PCRanalysesinenvironmentalmicrobiologyandevaluatethecontribu-Q-PCR;RT-Q-PCR;16SrRNAgene;mRNA.tionandadvancesgainedfromsuchappr
6、oaches.Finally,weconcludebyofferingfutureperspectivesontheapplicationof(RT)-Q-PCRinfurtheringunderstandinginmicrobialecology,inparticular,whencoupledwithothermolecularapproachesandmoretraditionalinvestigationsofenvironmentalsystems.aspyrosequencing(Marguliesetal.,2005;Edwardsetal.,Introduction2006)
7、nowdwarfPCR-basedsequencestudiesintermsofTheapplicationofPCRincombinationwiththeextractionsequencecoverage,theabilityofthePCRtospecificallyofnucleicacids(DNAandRNA)fromenvironmentaltargetparticulartaxonomico
