番茄果实乙烯信号转导组分EREBPs基因克隆和表达研究

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1、中国农业大学博士学位论文番茄果实乙烯信号转导组分EREBPs基因克隆和表达研究姓名：蔚变云申请学位级别：博士专业：食品科学指导教师：罗云波20030501AbstractTheplanthormoneethylenehasaprofoundroleinplantgrowthanddevelopment．Anumberofethylanesignalingelementshavebeenisolatedandidentified．Althou【ghnuclearcascadesinethylenesignaltransductionpathwayfromE

2、IN3toERFltoPRproteinshasbeenestablishedinArabidopsisthaliana，asimilarroutefromEIN3toEREBPstofruitripeningandsenescencehasnotbeenelucidatedinLycopersiconesculentum(tomato)．EREBPs(ethylene—responsiveelementbindingproteins)belongtoafamilyoftranscriptionfactors，whichregulateethylener

3、esponsesthroughbindingtothepromoterregionofethylene—regulatedtargetgenes．StudiesonEREBPsintomato，whichisamodelplantinfruitandvegetableripeningandsenescenceresearches，willmakeusbeRerunderstandthemechanismofripening-relatedgenesexpressionregulatedbyethyleneandfurtherelucidateethyle

4、netransductionpathway．Apairofdegeneraleprimersweredesignedintheconserveddomain(ERFdomain)whichbasedonthealignmentofEREBPsgeaefamilyintomatoandotherplantspecies．A14bpfragmentwasamplifiedbyRT—PCR(ReverseTranscriptionPolymeraseChainReaction)，whichwasusedasaprobeforscreeningtomatofru

5、it(pinkstage)cDNAlibrary．Twopositiveclonescontainingentirecodingregionwereisolated，whichwereidentifiedasnewmembersofEREBPsfamilybyBI—ASTServer,andnamedasLeERFlandLeERF2．TheLeERFlcDNAis1041bpandthepredictedtranslationproductis204aminoacidsTheLeERF2cDNAis2898bpandthepredictedtransl

6、ationproductis210aminoacidsLeERFlwashomologonswithEREBP4andDDTFRl0A，with33％similarityattheaminoacidlevel．LeERF2washomologouswithEREBP3andpti5，with46％similarityattheaminoacidlevel．ThecDNAsequencesofLeERFlandLeERF2weredepositedinGenBank(GenBankAccessionNumberAY077626，AY275554respec

7、tively)．NorthemblothybridizationshowedthatLeERFl，LeERF2andpti4(amemberofEREBPsintomato)expresseddifferentiallyindifl'erentpartsofwildtomatofruitduringripening，whichwasrelatedwithfruitripening．InbothNrfruitandantisenseACStransgenictomatofruit，expressionlevelsofLeERFI，LeERF2andpti4

8、weregenerallylowerthanthoseinwildfruitbu