ITS序列分析在真菌分类鉴定中的应用陈剑山

ITS陈剑山1,2,郑服丛1,2*(1.,571737;2.,571737)综述了ITS序列分析技术的原理特点及近年来在病原真菌分类鉴定上的应用,同时对ITS序列分析技术在真菌分类鉴定研究领域中的前景进行了展望ITS;分类;鉴定Q789A0517-6611(2007)13-03785-02ApplicationofITSSequencesinFungiClassificationandIdentificationCHENJian-shanetal(EnvironmentandPlantProtectionCollege,SCUTA,Danzhou,Hainan571737)AbstractTheprinciple,characteristicsandtheapplicationofITSsequencesinfungiclassificationandidentificationinrecentyearsweresummarized.Anditsprospectwasputforward.KeywordsITS;Classification;Identification,,,,1953WatsonCrickDNA,,(RestrictionFragmentLengthPolymorphism,RFLP)DNA(Am-plifiedFragmentLengthPolymorphism,AFLP)rDNA(InternalTranscribedSpacer,ITS)DNA(RandomAmplifiedPolymorphicDNA,RAPD)[1]PCRITS(InternalTranscribedSpacer)1ITSRNA(rRNA)55.817~18(18S)25~28S(28S),rRNA(rDNA)(5cy3c):¹(NonTranscribedSe-quence),NTS;º(ExternalTranscribedSpac-er),ETS;»18SrRNA,18SrDNA;¼1(InternalTranscribedSpacer1),ITS1;½5.8SrRNA,5.8SrDNA;¾2,ITS2;¿28SrRNA,28SrDNAITS1ITS2ITS,5.8SRNAITS[2]2,ITS1ITS4ITS(18S)ITS15.8SITS2(28S)(1),,60~200陈剑山(1982-),男,福建莆田人,硕士研究生,研究方向:分子植物病理学*通讯作者2006-12-21ITS650~750bp()[3]1ITS2ITSrDNA5.81828SrRNA,ITS,ITS,,2ITS,[4],ITS18SrDNA10ITS[5]ITS1ITS2,,ITSITS,ITS,YanITSPhialopho-raamericanaP.verrucose,[6](Tilletiaindica)ITS,(T.walkeri)T.indica,[7]ITSLentinulaSuillusTuberCeratocystisOidiodendron,[8-10]DNA-ITS,GenBankITS99.87%,1,(Phytophthoracrypto-gea)[11]DNAITS,298.28%,,JournalofAnhuiAgri.Sci.2007,35(13):3785-3786,3792刘月娟胡先祥99.17%,,,,[12],ITSGenBankITS,,[13]3ITSITS,,,ITSrDNAITS(PCR)rDNA,DNAITSrDNA,PCRrDNAPCR[14]:¹DNA,0.1~10ng;ºDNA2,;»DNA;¼PCR,WhiterRNAITS3,ITS1ITS4ITS5,[15]ITSITS1-FITS4-BGardes[16](1),Marmeisse(Hebeloma)[17]KamelITS[18]ITS1ITS218SrD-NA5.8SrDNAITS1,ITS3ITS45.8SrDNA28SrDNAITS2[19]1ITS(5.y3.)ITS1TCCGTAGGTGAACCTGCGC18SITS1-FCTTGGTCATTTTAGAGGAAGTAA18SITS1-R(TA)TGGT(CT)(AGT)(TC)(TC)TAGAGGAAGTAA18SITS5GGAAGGTAAAAGTCAAGG18SITS4TCCTCCGCTTATTGATATGC28SITS4-BCAGGAGACTTGTACACGGTCCAG28SITS4-RCAGACTT(GA)TA(CT)ATGGTCCAG28S,ITS,,,ITSITS,,,,18SITS,18S,ITSITS,,,,ITS,,,,18S28SrRNA,,,ITS,ITS,,[20]4,,DNA,,ITSITSITS,,,,DNA,,ITS[1]赵瑞琳.RAPD在真菌分类鉴定上的应用[J].热带农业科技,2004,27(3):23-26.[2]林晓民,李振岐,王少先.真菌rDNA的特点及在外生菌根菌鉴定中的应用[J].西北农业学报,2005,14(2):120-125.[3]赵杰.ITS序列分析及其在植物真菌病害分子检测中的应用[J].陕西农业科学,2004(4):35-37.[4]郑雪松,杨虹,李道棠,等.基因间隔序列(ITS)在细菌分类鉴定和种群分析中的应用[J].应用与环境生物学报,2003,9(6):678-684.[5]LEBLOND-BOURGETN,HERVEP,IRENEM,etal.16SrRNAand16Sto23Sinternaltranscribedspacersequenceanalysesrevealinterandintraspecificbifidobacteriumphylogeny[J].IntJSystBacteriol,1996,46:102-111.[6]YUANGF,LIUCS,CHENCC.DifferentiationofAspergillusparasiticusfromAspergillussojaebyRandomamplificationofpolymorphicDNA[J].Ap-pliedandEnvironmentalMicrobiology.1995,61(6):2384-2387.[7]章桂明.小麦印度腥黑粉菌及其近似种的分子系统发育分析和形态学比较[D].华南农业大学,1999.[8]HIBBERTDS,FUKUMASANAKAIY,TSUNEDAA,etal.Phylogeneticd-iversityinshiitakeinferredfromnuclearribosomalDNAsequences[J].Mycolo-gia.1995,87(5):618-638.[9]GANDEBOEUFD,DUPREC,DREVETP,etal.GroupingandidentificationofTuberspeciesusingRAPDmarkers[J].CanJBot,1997,75(1):36-45.[10]HAMBLETONS,EGGERKN,CURRAHRS.ThegenusOidiodendron:speciesdelimitationandphylogeneticrelationshipsbasedonnuclearribosomalDNAanalysis[J].Mycologia,1998,90(5):851-869.[11]张正光,郭成宝,王源超,等.非洲菊根腐病病原的鉴定与ITS序列分析[J].植物病理学报,2005,35(5):392-396.[12]杨雅云,罗文富,杨艳丽.马铃薯及番茄晚疫病菌的核糖体DNA-ITS(下转第3792页)3786安徽农业科学2007,0.01mol/LPBS,(200~340nm)12,2,4-D-BSA2,4-D-OVA214.9210.1nm279.7268.2nm2,4-DBSAOVA,,2,4-DBSAOVA,2,4-D-BSA2,4-D-OVA[9]12,4-D2,4-D-BSABSA22,4-D2,4-D-OVAOVA,2:¹;º,2,4-D,1-OCH2COOH,2.2,BSA2,4-D24B1,OVA2,4-D17B1,[6],,,,,(34~5)B1,[10];,(10~20)B1[6],,(15~25)B1,2.3,ELISA2,4-D,A1.02,4-D-OVA4.0Lg/ml,2.56@1041.28@1041.28@104ELISA2,4-D-OVA1.0Lg/ml,0.1Lg/ml32,4-D(2,4-D-BSA)(2,4-D-OVA),ELISA2,4-D,22,4-D2,4-D[1]李金昶,王璐,韩明友,等.固相萃取富集高效液相色谱法测定苯氧乙酸和2,4-二氯苯氧乙酸[J].分析化学,2001,29(5):580-582.[2]左凤,夏敏,谷学新.高效液相色谱法检测两种苯氧乙酸类除草剂[J].现代科学仪器,2003(4):63-64.[3]CUONGNV,BACHMANNTT,SCHMIDRD.Developmentofadipstickim-munoassayforquantitativedeterminationof2,4-dichlorophenoxyaceticacidinwater,fruitandurinesamples[J].FreseniusJAnalChem,1999,364:584-589.[4]FRANEKM,KOLARV,GRANATOVAM,etal.MonoclonalELISAfor2,4-dichloroPhenoxyaceticacid:Characterizationofantibodiesandassayoptioization[J].JAgricFoodChem,1994,42:1369-1374.[5]MATUSZCZYKG,KNOPPD,NIEBNERR.DevelopmentofanELISAfor2,4-D:characterizationoftwopolyclonalantisera.Fresenius[J].JAnalChem,1996,354:41-47.[6]陈建新,陈梅英,赵会杰.免疫学技术在植物科学中的应用[M].北京:中国农业大学出版社,1998:54-58.[7]邓瑞春.两种不同方法纯化抗血清IgG的效果比较[J].免疫学杂志,1999,15(1):64-66.[8]薛小平,张美顺,李荣源,等.高亲和力抗有机磷杀虫剂单克隆抗体的制备与鉴定[J].中国生物工程杂志,2003,23(6):68-71.[9]孙远明,赵肃清,张春艳,等.甲胺磷人工抗原的合成与鉴定[J].免疫学杂志,2002,18(3):163-166.[10]吴颂如,万寅生,周燮.植物小分子物质的免疫测定技术[J].植物生理通讯,1989,5:68-72.(上接第3786页)区段序列分析[J].云南农业大学学报,2005,20(2):188-192.[13]杨腊英,黄华平,唐复润,等.香蕉炭疽菌rDNAITS区的分子鉴定与检测[J].植物病理学报,2006,36(3):219-225.[14]张志华,洪葵.核酸序列直接分析在真菌鉴定方面的应用[J].华南热带农业大学学报,2006,12(2):39-43.[15]WHITETJ,BRUNST,LEES.Analysisofphytogeneticrelationshipsbyam-plificationanddirectsequencingofribosomalRNAgenes[C]//INNISMA.PCRProtocols:AGuidetoMethodsandApplications.NewYorkAcademic,1990,315-322.[16]GARDESM,BRUNSTD.ITSprimerwithenhancedspecifi