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1、MINIREVIEWAdvantagesandlimitationsofquantitativePCR(Q-PCR)-basedapproachesinmicrobialecologyCindyJ.Smith&A.MarkOsbornDepartmentofAnimalandPlantSciences,UniversityofSheffield,WesternBank,Sheffield,UKCorrespondence:A.MarkOsborn,AbstractDepartmentofAnimalandPlantSci
2、ences,QuantitativePCR(Q-PCRorreal-timePCR)approachesarenowwidelyappliedUniversityofSheffield,WesternBank,SheffieldS102TN,UK.Tel.:144114222inmicrobialecologytoquantifytheabundanceandexpressionoftaxonomicand4626;fax:1441142220002;e-mail:functionalgenemarkerswithint
3、heenvironment.Q-PCR-basedanalysescombinea.m.osborn@sheffield.ac.uktraditionalend-pointdetectionPCRwithfluorescentdetectiontechnologiestorecordtheaccumulationofampliconsinrealtimeduringeachcycleofthePCRReceived20June2008;revised10Octoberamplification.Bydetectionofa
4、mpliconsduringtheearlyexponentialphaseofthe2008;accepted24October2008.PCR,thisenablesthequantificationofgene(ortranscript)numberswhentheseFirstpublishedonlineDecember2008.areproportionaltothestartingtemplateconcentration.WhenQ-PCRiscoupledwithaprecedingreversetr
5、anscriptionreaction,itcanbeusedtoquantifygeneDOI:10.1111/j.1574-6941.2008.00629.xexpression(RT-Q-PCR).ThisreviewfirstlyaddressesthetheoreticalandpracticalimplementationofQ-PCRandRT-Q-PCRprotocolsinmicrobialecology,high-Editor:MichaelWagnerlightingkeyexperimental
6、considerations.Secondly,wereviewtheapplicationsofKeywords(RT)-Q-PCRanalysesinenvironmentalmicrobiologyandevaluatethecontribu-Q-PCR;RT-Q-PCR;16SrRNAgene;mRNA.tionandadvancesgainedfromsuchapproaches.Finally,weconcludebyofferingfutureperspectivesontheapplicationof
7、(RT)-Q-PCRinfurtheringunderstandinginmicrobialecology,inparticular,whencoupledwithothermolecularapproachesandmoretraditionalinvestigationsofenvironmentalsystems.aspyrosequencing(Marguliesetal.,2005;Edwardsetal.,Introduction2006)nowdwarfPCR-basedsequencestudiesi
8、ntermsofTheapplicationofPCRincombinationwiththeextractionsequencecoverage,theabilityofthePCRtospecificallyofnucleicacids(DNAandRNA)fromenvironmentaltargetparticulartaxonomico
