英文文献阅读报告课件.ppt

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1、ReadingReportAgrobacterium农杆菌是生活在植物根的表面依靠由根组织渗透出来的营养物质生存的一类普遍存在于土壤中的革兰氏阴性细菌。农杆菌是一种天然的植物遗传转化体系，被誉为“自然界最小的遗传工程师”。可以通过将目的基因插入到经过改造的T-DNA区，借助农杆菌的感染实现外源基因向植物细胞的转移和整合，然后通过细胞和组织培养技术，得到转基因植物。Agrobacteriumrhizogenes-mediatedtransformationprovidesasystemforrapidandefficie

2、nttransformationofplanttissues.TheinfluenceofseveralfactorsplantgenotypeA.rhizogenesculturestageA.rhizogenescelltitreco-cultureperiodofA.rhizogenesacetosyringoneconcentrationaroutine,efficientandgenotype-independentmethodofinducinghairyrootsbyA.rhizogenesinpeanut

3、swasestablished.诱导农杆菌Vir基因活化，从而促进外源基因的整合PlantmaterialSevenpeanutgenotypesincludingLuhua11,Huayu16,Huayu28,Baisha1016,Yuanhua8,Xinhua1,andXinhua5wereusedtoassesstheeffectofplantgenotypeonA.rhizogenes-mediatedtransformation.Agrobacteriumrhizogenesstrainandplasmid

4、pCAMBIA2300ThevectorpGFP-GUSPluswhichcontainedtwohighly-expressedreportergenes,GFPgeneandGUSgene.InductionofrootsbymicroinjectionFig.1Peanuthairy-rootinductionA-5-dayoldseedlingformicroinjection;B-microinjectionofA.rhizogenesonhypocotylsatdifferentsides;C-Tumor

5、(肿瘤)emerging1weekaftermicroinjection;D-Inducedrootsappearing2weeksafterinoculation;E-TransformedrootsystemDetectionofGFPorsyntheticcry8Ea1geneintransgenicrootsbyRT-PCRTotalRNAfrominducedrootswasisolatedbytheTRIzolmethodandtreatedwithRNase-freeDNaseaccordingtoth

6、emanufacturer’sinstructions.OnemicroliteroffirststrandcDNAwasusedastemplateforPCRamplification.ThetwosetsofprimerpairsusedintheassaywereprimersVirHF/VirHR,primerGFPF(basepairs76–93),50-CTTCTCGTTGGGGTCTTT-30,andprimerGFPR(basepairs627–644),50-ACAAGTTCAGCGTGTCCG-30

7、.Primerjc8EF2,50-GGCGGCAGCATTCAAACTCAA-30,andprimerjc8ER2,50ATCTCCACCAAGATAGTGTCC-30wereusedtodetectthetranscriptionofthesyntheticcry8Ea1gene（暗黑鳃金龟抗性基因）.HistochemicalassayforGUSactivityRootsegments(2cmlong)andhypocotylswerehistochemi-callystainedtoanalyzeGUSgen

8、eexpressionaccordingtoJefferson.Fig.2GUSstainingassayGUSexpressioninverticalsectionVerticalsectiontransversesectiontransversesectionhypocotylsinjectedwithK599andinducedroots