Development of Useful Recombinant Promoter and Its Expression Analysis in Different Plant Cells Using Confocal Laser Scanning Microscopy 英文参考文献.docVIP

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Development of Useful Recombinant Promoter and Its Expression Analysis in Different Plant Cells Using Confocal Laser Scanning Microscopy 英文参考文献.doc

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Development of Useful Recombinant Promoter and Its Expression Analysis in Different Plant Cells Using Confocal Laser Scanning Microscopy 英文参考文献

DevelopmentofUsefulRecombinantPromoterandIts ExpressionAnalysisinDifferentPlantCellsUsing ConfocalLaserScanningMicroscopy DeepakKumar1,SunitaPatro1.,RajivRanjan1.,DipakK.Sahoo1,2,InduB.Maiti2,NrisinghaDey1* 1Department of Gene Function and Regulation, Institute of Life Sciences, Department of Biotechnology, Government of India, Nalco Square, Chandrasekherpur, Bhubaneswar,Orissa,India,2KentuckyTobaccoResearchandDevelopmentCenter(KTRDC),CollegeofAgriculture,UniversityofKentucky,Lexington,Kentucky,United StatesofAmerica Abstract Background:Designingfunctionallyefficientrecombinantpromotershavingreducedsequencehomologyandenhanced promoteractivitywillbeanimportantsteptowardsuccessfulstackingorpyramidingofgenesinaplantcellfordeveloping transgenicplantsexpressingdesiredtraits(s).Alsobasicknowledgeregardingplantcellspecificexpressionofatransgene undercontrolofapromoteriscrucialtoassessthepromoter’sefficacy. Methodology/Principal Findings: We have constructed a set of 10 recombinant promoters incorporating different up- streamactivationsequences(UAS)ofMirabilismosaicvirussub-genomictranscript(MS8,-306to+27)andTATAcontaining core domains of Figwort mosaic virus sub-genomic transcript promoter (FS3, 2271 to +31). Efficacies of recombinant promoterscoupledtoGUSandGFPreportergenesweretestedintobaccoprotoplasts.Amongthese,a369-bplonghybrid sub-genomictranscriptpromoter (MSgt-FSgt)showedthehighestactivityinbothtransientandtransgenicsystems.Ina transientsystem,MSgt-FSgtwas10.31,2.86and2.18timesmoreactivecomparedtotheCaMV35S,MS8andFS3promoters, respectively. In transgenic tobacco (Nicotiana tabaccum, var. Samsun NN) and Arabidopsis plants, the MSgt-FSgt hybrid promoter showed 14.22 and 7.16 times stronger activity compared to CaMV35S promoter respectively. The correlation betweenGUSactivityanduidA-mRNAlevelsintransgenictobaccoplantswereidentifiedbyqRT-PCR.BothCaMV35Sand MSgt-FSgt promoters caused gene silencing but the degree of silencing are less in the case of the MSgt-FSgt pro