16SrRNA基因技术在油藏微生物生态研究中的应用

:10050930(2007)030369011:TE3579;Q9381:A:20060224;:20070529:(No.50604013);(2006BAB04B02);(2003BA613A08).:(1971!),,.Emai:[e-mail]16SrRNA包木太1,肖生科2,孔祥平1,郭省学3,李希明3,李阳4,王修林1(1.,266100;2.,735200;3.,257000;4.,100029):分子生态学技术的发展大大促进了人们对环境中微生物群落的研究,也为研究油藏微生物群落和微生物采油技术提供了一个新的工具,尤其16SrRNA基因技术应用于油藏微生物生态研究.该技术克服了基于细胞水平研究方法的不足,能更准确更客观地揭示油藏微生物的多样性群落动态变化种群结构及进化等方面的信息.文章简要综述了16SrRNA基因技术发展历程及在环境微生物生态学中的应用,详细概述了16SrRNA基因技术在油藏微生物生态研究中的地位和作用以及用于油藏微生物生态研究的16SrRNA基因技术,讨论了16SrRNA基因技术目前在油藏微生物生态研究中存在的问题,通过实例论证了该技术的现场应用效果,重点介绍了胜利油田利用TRFLPDGGE等分子生态学技术在河口采油厂罗801区块空气辅助微生物驱和单12区块内源微生物驱研究中的应用,阐明了分子生态学技术在微生物提高原油采收率研究中的重要的意义.:16SrRNA;;;,2l[12].,.,.,.,(50!130∀)(10!50MPa)///,.15320079JOURNALOFBASICSCIENCEANDENGINEERINGVo.l15,No.3September2007,,[3],(HOB)(FB)(DNB)(TGB)(SRB)(MPB)[4].,[5],(viablebutnonculturable,VBNC),01%!10%[68],.,,,,[9].,,[10].,,..116SrRNArRNA,rRNA,rRNA5#16S23S5S3#[11],5S16S23SrRNA[12].rRNA,,,[13].16SrRNA16SrDNA,,[14].,,1500bp,16SrRNA,,16SrRNA[15].16SrRNA,,[16].,16SrRNA(PolymeraseChainReaction,PCR),.16SrRNA,.,16SrRNA16SrRNA,.1986Pace[17],,,16SrRNA,,.DGGEFischerLermanDNA370Vo.l15,1993Muyzer[18],,.2004[19]16SrDNA(ribosomalintergenicspaceranalysis,RISA).[20]16SrRNA.Janus[21]16SrRNACO2.[22]816SrRNA..216SrRNA,,[2326],,16SrRNA,,.[27]RFLP16SrRNA.[28](PolymeraseChainReactionTemperatureGradientGelElectrophoresis,PCRTGGE),.DNA,;RNA,,,.21,.,.22,.16SrRNA,,.23,,,,01%!10%.,16SrRNA,.2416SrRNA,DGGE371No.3:16SrRNA,,[2930].25,,.,,,.316SrRNA16SrRNA16SrRNA(DenaturingGradientGelElectrophoresis,DGGE)(RestrictionFragmentLengthPolymorphism,RFLP)(SingleStrandConformationPolymorphism,SSCP)(fluorescentinsituhybridization,FISH),.DGGE.3116SrRNA16SrRNA1500bp,[31],16SrRNA,GeneBank,,.,16SrRNA[32].DNA,,PCRDNA16SrRNA.,,,,.32(DGGE)DNA,PCRDNA,DGGE.DGGE,DGGE,[7].3.2.1DGGEDGGE,DNA.DNA(),,DNA,,,DNA,DNA,,[33].3.2.2DGGE(1);(2);(3)DNA;(4)DNA16SrRNA;(5)DGGE;(6)DGGE.372Vo.l1533(RFLP)(RFLP)DNA,DNA[34].,.(TerminalRestrictionFragmentLengthPolymorphism,TRFLP)RFLP,,,.PCR,,[35].LiuTRFLP,[36].RsaI,MspI16SrRNAPCR,.RDP.34(SSCP)(SSCP).DNA,.,,[35,37].DNA.,,.PCR16SrRNA,SSCP,SSCP.35,,DNAPCR.,,,,[38].:(1);(2);(3)16SrRNA;(4)rRNA;(5);(6).416SrRNA41DNA.,DNA[6].,DNA16SrRNA373No.3:16SrRNA,,.42PCR4.2.1TaqPCRTaq,Taq,16SrRNA.4.2.2PCRPCR,.4.2.3PCR(),16SrRNA,PCR[39].43,,,,rRNA[10].4416SrRNADGGERFLP..516SrRNA51PCRDNA,.:,[23].52PCRPCRDGGE,16SrRNAPCRDGGEDNA,(12911)(1299S13118).DGGE30%20%,54%.DGGE,!(Proteobacteria)(Bacteroidetes).DGGE(MEOR),MEOR[40].53TRFLP374Vo.l15801,2004TRFLP.:801,,50%,.,[41].TRFLP16SrDNA,(RDP),,.,,.,,.54PCRDGGETRFLP,PCRDGGETRFLP12.DGGE:12,,;TRFLP,12,.,[42].,PCRDGGE[43].616SrRNA,,.16SrRNA,.,,,,.,,[44].,16SrRNADNAPCR,,.375No.3:16SrRNA[1],,,.[J].,2002,19(4):382386BaoMuta,iWangWeidong,WangXiulin,eta.lMicrobialenhancedoilrecoverybyactivationofstratalmicroflora[J].OilfieldChemistry,2002,19(4):382386[2],,,.[J].,2000,8(3):236245BaoMuta,iMuBozhong,WangXiulin,eta.lMicrobialenhancedoilrecovery[J].JournalofBasicScienceandEngineering,2000,8(3):236245[3].[J].,1995,22(6):383384;357WangXiuyuan.TheadvancementofmicrobialenhancedoilrecoveryinRussia[J].Microbiology,1995,22(6):383384;357[4],,,.[J].,2003,20(4):372376KongXiangping,BaoMuta,iMaDaixin,eta.lAnalysisofbacterialcommunitiesinoilfieldproducedwaters[J].OilfieldChemistry,2003,20(4):372376[5]JyhYihLeu,CarolinePMcGovernTraa,AndrewJRPorter,eta.lIdentificationandphylogeneticanalysisofthermophilicsulfatereducingbacteriainoilfieldsamplesby16SrDNAgenecloningandsequencing[J].Anaerobe,1998,4:165174[6]BrockTD.Thestudyofmicroorganismsinsitu:progressandproblems[J].Symp.Soc.Gene.Microbio.l,1987,41:117[7]AmannRI,LudwigW,SchleiferKH.Phylogeneticidentificationandinsitudetectionofindividualmicrobialcellswithoutcultivation[J].Microbio.lRev.,1995,59(1):143169[8]McCaigAE,GloverLA,ProsserJI.Molecularanalysisofbacterialcommunitystructureanddiversityinunimprovedandimproveduplandngrasspastures[J].ApplEnvironMicrobio.l,1999,65(4):17211730[9]DunbarJ,WhiteS,ForneyL.Geneticdiversitythroughthelookingglass:effectofenrichmentbias[J].App.lEnviron.Microbio.l,1997,63:13261333[10],,,.rRNA[J].,2001,21(1):3741HuangLinan,QuLianghu,ChenYueqin,eta.lrRNAtechniqueanditsapplicationinmolecularmicrobialecology[J].JournalofMicrobiology,2001,21(1):3741[11],,,.16SrRNA[J].,2001,22(5):265268LiuJiawen,LvHongyan,WuXueqiong,eta.lThedevelopmentof16SrRNAgenesequenceinsortidentifyofmycobacterium[J].FasciculeofForeignMedicalClinicBiochemistryandTest,2001,22(5):265268[12]SchmidtTM,DelongEF,PaceNR.Analysisofamarinepicoplanktoncommunityby16SrRNAgenecloningandsequencing[J].Bacterio,l1991,173(14):43714378[13],,,.16SrRNA[J].,1998,25(1):1216JiaoZhenquan,LiuXiume,iMengZhaohe,eta.lThehomologyanalyzationof16SrRNAsequenceandthesortidentificationofbacteriumsystem[J].FasciculeofForeignMedicalHygienics,1998,25(1):1216[14]CelineDelbes,JeanJacquesGodon,ReneMoletta.16SrRNAsequencediversityofacultureaccessiblepartofananaerobicdigestorbacterialcommunity[J].Anaerobe,1998,49(4):267275[15]FerrisMJ,WardDM.Seasonaldistributionsofdominant16SrRNAdefinedpopulationsinahotspringmicrobialmatexaminedbydenaturinggradi