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1、Cloning,expressionanalysis,anddetectionmethodoftheS12-RNasegeneexistinginthreepearspecies(Pyrusbretschneideri,P.pyrifolia,andP・ussuriensis)ZhangLim,LiuMini,TanXiaofengi,SongZhiboi,LongHongxui,CaoYufem,LiXiugen3(1.KeyLaboratoryofCultivationandProtect!onforNon-WoodForestTrees,MinistryofEducation,Cent
2、ralSouthUniversityofForestryandTechnology,ChangSha410004;2.ResearchInstituteofPomology,ChineseAcademyofAgriculturalSciences,LiaoNingXingCheng125100;3.ZhengzhouFruitResearchInstitute,ChineseAcademyofAgriculturalSciences,ZhengZhou450009)Abstract:Aspecificforwardprimerwasdesignedbasedontheprincipleofh
3、ighestsimilarity,andemployedin3'RACEusingChinesewhitepear(Pyrusbretschneideri)cultivar'Yingzhiqing'asmaterials.ThefulllengthcDNAofPbS12-RNascwassuccessfullyamplifiedanddepositedinGenBank(AccessionNo.EU081889).Attheaminoacidlevel,thePbS12-RNaseexhibitedthehighestsimilarity(97.3%)withMdSf-RNascofMalu
4、sdomcstica,andonlysixaminoaciddifferenceswerepresentinthetwoS-RNascs.PhylogeneticanalysisofrocaccousS-RNascsindicatedthatthePbS12-RNascclusteredwithmaloidcousS-RNascs,formingasubfamily-specificnotspccics-spccificgroup・SomeintraspecificgeneticdistanceoftheS-RNascs,inaddition,wasgreaterthaninterspeci
5、ficgeneticdistance・In'YingzhiqingthePbS12-allelewasspecificallyexpressedinstyle.Moreover,theexpressionlevelofthisgenewasextremelylowatthesmallbudstage,andsubsequentlyincreasedrapidlyatthebidbudstageandbellstage・AmethodforrapiddetectionofthePbS12-allelewasdevelopedviaPbS12-allele-speceficprimersdes
6、ignbasedonmultiplesequencecomparisons.ApplicationofthePbS12-allele-speceficprimersin59cultivarsfromfourpearspeciesshowedthatthePbS12-allelenotonlyexistedinP.bretschneideri,butinP.pyrifoliaandP.ussuriensisaswell.ThepresentstudycouldprovideascientificbaseforfullyclarifyingthemechanismofpearGSIatthemo
7、lecularlevel.Keywords:ganietophyticself-incompatibility;reversetranscript-PCR;RapidAmplificationofcDNAEnds;S-RNasc;phylogenetictree;spatio-temporalexpressionpatterns0IntroductionSelf-incompatibility(SI)isan
