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1、中國農(nóng)業(yè)大學博士學位論文番茄果實乙烯信號轉(zhuǎn)導組分EREBPs基因克隆和表達研究姓名:蔚變云申請學位級別:博士專業(yè):食品科學指導教師:羅云波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