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1、萬(wàn)方數(shù)據(jù)作物學(xué)報(bào)ACTAAGRONOMICASINICA2011,37(8):1389--1397http://www.chinaerops.org/zwxb/ISSN0496-3490;CODENTSHPA9E—mail:xbzw@chinajoumal.net.CUDoI:10.3724/SP.J.1006.2011.01389小麥TaCRF2基因的克隆及其在煙草中的初步功能驗(yàn)證郭麗香1,2趙昌平2·’高世慶2,料唐益苗2王永波2劉美英3張朝3徐蓓1,21首都師范大學(xué)生命科學(xué)學(xué)院,北京100048;2北京市農(nóng)林科學(xué)院雜交小麥
2、工程技術(shù)研究中心.北京100097;3中國(guó)科學(xué)技術(shù)大學(xué)研究生院化學(xué)系,安徽合肥230026摘要:通過(guò)RT-PCR方法從小麥cDNA中擴(kuò)增獲得1個(gè)鋅指蛋白基因TaCRF2,該基因的cDNA長(zhǎng)度為843bp,序列分析表明它編碼1個(gè)含有280個(gè)氨基酸的蛋白質(zhì)。在線軟件預(yù)測(cè)該蛋白質(zhì)的相對(duì)分子質(zhì)量為30.97kD,等電點(diǎn)為7.03,且在C一末端含有1個(gè)典型的RING.H2型鋅指蛋白結(jié)構(gòu)域,在N.末端含有2個(gè)跨膜結(jié)構(gòu)域。氨基酸序列比對(duì)發(fā)現(xiàn),TaCRF2與水稻中1個(gè)RING型鋅指蛋[芻(ABF95226)的相似度為82%。亞細(xì)胞定位分析顯示,
3、該蛋白分布在細(xì)胞核和細(xì)胞膜上。real.timeRT-PCR表達(dá)特性分析顯示,TaCRF2基因的表達(dá)受干旱、高鹽和外源ABA的強(qiáng)烈誘導(dǎo),低溫對(duì)該基因的表達(dá)量影響不明顯。初步功能驗(yàn)證發(fā)現(xiàn)過(guò)表達(dá)TaCRF2基因增強(qiáng)了轉(zhuǎn)基因煙草對(duì)干旱和高鹽脅迫的耐性。關(guān)鍵詞:煙草;小麥;RING.H2型鋅指蛋白;TaCRF2基因;干旱脅迫;鹽脅迫TaCRF2GeneIsolationfromTriticumaestivumandPrimaryFunctionValida-tioninTobacco(NicotianatabacumL)GUOLi—Xi
4、an91,一,GAOShi.Qin92’”,TANGYi.Mia02,WANGYong.B02,L1UMei—Yin93,ZHANGZha03,XUBcilt_.a(chǎn)ndZHAOChang.Pin92'‘1CollegeofLifeSciences.CapitalNormalUniversity,Beijing100048,China;2BeijingHybridWheatEngineeringandTechnologyResearchCenter,BeijingAcademyofAgricultureandForestrySci
5、ences,Beijing100097,Chhla;3DepartmentofChemistry,GraduateSchoolofUniversityofScienceandTechnologyofChina,Hefei230026,ChinaAbstract:ⅥmeaLlikeothercrops。suffersfromdrought,salt,low.temperatureandotherabioticstresses,seriouslyresultinginyielddecline.ItisaneffectiveWayfo
6、rcropbreedingtoimproveresistanceviamolecularbiologytechniques.Zincfmgerprotein,animportanttranscriptionfactorcommonlyfoundinplant,carlregulatetheexpressionofmultiplestress—induciblegenesandenhancecomprehensiveresistanceseffectively.Inthisstudy,aIu時(shí)G-H2zincfingerprote
7、ingene,designatedasTaCRF2,wasisolatedfromTriticumaestivumbvRT-PCR.ItscDNAwas843bpandencodedaputativeproteinof280Rlllinoacidswithapre-dictedmolecularmassof30.97kDandanisoelectricpoint(PI)of7.03.AtypicalRENG.H2fingerdomainwasfoundattheC-terrainalregionofT忙RF2protein.a(chǎn)n
8、dtWotransmembranedomainswerefoundattheN-terminalregion.AlignmentofaminoacidsequenceshowedthatTaCRF2was82%identicaltoOryzasativaputa