嘉南药理科技大学环境工程与科学系硕士论文

IAnalysisoftheChangesofPhenolDegradersbyDenaturingGradientGelElectrophoresisDuringPhenolDegradationinsoilsIIDepartmentofEnvironmentalEngineeringandScience.Chia-NanUniversityofPharmacyandScienceThesisfortheDegreeofMasterAnalysisoftheChangesofPhenolDegradersbyDenaturingGradientGelElectrophoresisDuringPhenolDegradationinsoils(Dr.Rey-MayLiou)(Dr.Shuh-RenJing)(Ju-YuYang)30,June2005IIIIVV2DNA2VIM1M5M1M51,2-16SrDNAM1Ralstoniabasilensis(AB109778)96%M5Staphylococcussp.B60(AF333342)94%DNA341GC+534rPCRVIIAbstractPhenolisacommonpollutantfromindustries,itimpactsontheaquaticecosystemandenvironmentalbalanceextremely,andbiodegradationisaveryeconomictechnique,whichisthebestchoiceofthephenolcontaminatedsoilremediation.Thephenolhigh-tolerancedegraderswereisolatedfromcollectedsludgeandsoilfrompetrolstationandaroundofreactorandwastewaterstorageinSyndyneIndustry,thetestedstrainsinthisstudyincludingM1andM5.ThephysiologicalpropertiesoftestedstrainswereanalyzedbyGramstain,activityofcatecholdioxygenase.TheresultsshowedthatthetestedstrainswereGrampositiveandhadtheactivitiesofcatechol1,2-dioxygenase.TheM1andM5wereshowed96%probabilityofRalstoniabasilensis(AB109778)andStaphylococcussp.B60(AF333342)analysedinthedenaturinggradientgelelectrophoresis(DGGE)profilesofamplified16SrDNAfragment.Inoculationofeffectivedegraderstothemodifiedpollutedsoilsspikedwithphenolwascarriedoutbybatchculture.Thephenolconcentrationandthenumberofphenoldegraderweremonitoredintheexperimentalperiod,soilDNAwasextractedandamplifiedbyprimer341GC+534rofPCRforDGGEanalysisatthesametime,whichtounderstandthevariationofteststrains.Theresultsshowedthatthebiodegradationinthepollutedsoilsandaquaticm-cresolsolutionwerenotconsistent.Allthestrainscandegradephenolinthetestpollutedsoilsefficiently,themosteffectivedegradationinthesoilcollectedfromwasobservedbyteststrains.TracingsoilbacterialcommunitybyDGGEtechnologyintheexperimentalperiodwasobservedthatinoculatedeffectivestrainsbecamedominant,andthisresultwassimilartothecolonyVIIIformingunit(CFU)wascountedbyplate-countmethods.Thedenaturinggradientgelelectrophoresis(DGGE)isconsideredoneofthemostsensitiveofthescanningtechniquesforsoilcommunity.IXIIIIVVIIIX................................................................................................11.1...........................................................................................11.2........................................................................2.........................................................................................42.1.........................................................................42.1.1...........................................................................42.1.2..........................................................................42.2........................................................................................82.2.1.................................................................................82.2.2..................................................................................82.3...................................................112.3.1(polymerasechainreaction;PCR).................112.3.2(fluorescencein-situhybridizationFISH).....12X2.3.3(restrictionfragmentlengthpolymorphismRFLP).............................................................................................122.3.4(denaturinggradientgelelectrophoresisDGGE)............................................................................................12.....................................................................................153.1..............................................................................153.2.....................................................................................153.3.........................................................................................153.4.........................................................................................183.5..................................................................................183.6......................................................................................193.7............................................................193.7.1.............................................................................193.7.2.............................................................................203.7.3.....................................................................203.816SrDNA................................................................................213.8.1DNA..........................................................................213.8.2..................................................223.8.316SrDNA.......................................................................223.9..........................................................................23XI3.9.1........................................................................233.9.2DNA.....................................................................233.9.3..............................