資源描述：
《小麥tawrky46-1基因克隆及建立在gateway技術(shù)上的可》由會員上傳分享，免費(fèi)在線閱讀，更多相關(guān)內(nèi)容在工程資料-天天文庫。
1、華北農(nóng)學(xué)報2011,26(2):17-22小麥TaWRKY46-1基因克隆及建立在Gateway技術(shù)上的可誘導(dǎo)表達(dá)載體的構(gòu)建11111211李明,丁博,牛有雄,呂芳芳,楊文麗,SilinZhong,孫守鈞,謝曉東(1.天津農(nóng)學(xué)院農(nóng)學(xué)系,天津300384;2.BoyceThompsonInstituteforPlantResearch,CornellUniversity,NY14853,USA)摘要:以小麥品種揚(yáng)麥158為材料,根據(jù)NCBI中小麥TaWRKY46基因序列設(shè)計引物,采用RT-PCR技術(shù)從小麥總RNA中分離了大小為870bp的cDNA片段。測序表明,該片段
2、與NCBI中已克隆的小麥TaWRKY46基因高度同源,包含完整的讀碼框,值得注意的是此蛋白保守氨基酸序列WRKYGQK突變?yōu)閃RKYGEK,為TaWRKY46的新等位基因,命名為TaWRKY46-1。采用Gateway克隆技術(shù)構(gòu)建了該基因的可誘導(dǎo)型超量表達(dá)載體,并轉(zhuǎn)化農(nóng)桿菌,為接下來轉(zhuǎn)基因驗證TaWRKY46-1的功能奠定了基礎(chǔ)。關(guān)鍵詞:小麥;WRKY轉(zhuǎn)錄因子;Gateway克隆技術(shù);可誘導(dǎo)表達(dá)載體中圖分類號:Q785;S512.1035.3文獻(xiàn)標(biāo)識碼:A文章編號:1000-7091(2011)02-0017-06CloningofWheatTaWRKY46-1Gene
3、andConstructionofPlantInducibleExpressionVectorbyGatewayCloningTechnique11111LIMing,DINGBo,NIUYou-xiong,LUFang-fang,YANGWen-li,211SilinZhong,SUNShou-jun,XIEXiao-dong(1.TianjinAgriculturalUniversity,Tianjin300384,China;2.BoyceThompsonInstituteforPlantResearch,CornellUniversity,USA14853)Abst
4、ract:Inthisstudy,thewheatvariety,Yangmai158(TriticumaestivemL.),wasusedastheexperimentalma-terialforcloningthewheatTaWRKY46gene.PrimersweredesignedbasedonaknownwheatTaWRKY46genese-quenceinNCBI,subsequentlyan870bpcDNAfragmentwasamplifiedfromtotalRNAofYanmai158byRT-PCR.Sequenceanalysisrevealed
5、thattheclonedfragmentcontainsthefulL-lengthcDNAandishighlyhomologoustotheTaWRKY46sequenceinNCBI.ThesequencealignmentrevealedaQtoEchangewithintheconserved"WRKYGQK"domain,suggestingitwasanewalleleofTaWRKY46.HencewedesignateditasTaWRKY46-1.InordertoelucidatethefunctionalroleofthenewYangmai158Ta
6、WRKY46-1allele,aplantinducibleover-expressionvector,pKIGW-TaWRKY46-1,wasconstructedviaGatewaycloningtechnologyandtransformedintoAgrobacteriumcellssuccessfully.ThisworkprovidesthekeybasisforfuturedetailedfunctionalanalysisofwheatTaWRKY46-1gene.Keywords:Whea;tWRKYtranscriptionfactor;Gatewayclo
7、ningtechnology;PlantinducibleexpressionvectorWRKY基因家族是近年來新發(fā)現(xiàn)的鋅指型轉(zhuǎn)錄NCBI(NationalCenterforBiotechnologyInformation)調(diào)控因子,因在其N-端含有由WRKYGQK組成的高Gene數(shù)據(jù)庫中記錄的WRKY基因已達(dá)731條。度保守的7個氨基酸序列而得名。第一個WRKY轉(zhuǎn)WRKY結(jié)構(gòu)域及其專一結(jié)合的(T)(T)TGAC(C/T)[1]錄因子是由Ishiguro等于1994年從白薯中克隆序列(
