Agrobacterium rhizogenes-Mediated Transformation of the Parasitic Plant Phtheirospermum japonicum 英文参考文献.docVIP

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Agrobacterium rhizogenes-Mediated Transformation of the Parasitic Plant Phtheirospermum japonicum 英文参考文献.doc

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Agrobacterium rhizogenes-Mediated Transformation of the Parasitic Plant Phtheirospermum japonicum 英文参考文献

Agrobacteriumrhizogenes-MediatedTransformationof theParasiticPlantPhtheirospermumjaponicum JulianeK.Ishida1,2,SatokoYoshida2,MasakiIto3,ShigetouNamba1,KenShirasu2* 1Graduate School of Agricultural and Life Sciences, University of Tokyo, Tokyo, Japan, 2Plant Science Center, RIKEN, Yokohama, Japan, 3Graduate School of BioagriculturalSciences,NagoyaUniversity,Nagoya,Japan Abstract Background:PlantswithintheOrobanchaceaeareanagriculturallyimportantgroupofparasitesthatattackeconomically important crops to obtain water and nutrients from their hosts. Despite their agricultural importance, molecular mechanismsoftheparasitismarepoorlyunderstood. Methodology/Principal Findings: We developed transient and stable transformation systems for Phtheirospermum japonicum, a facultative parasitic plant in the Orobanchaceae. The transformation protocol was established by a combination of sonication and acetosyringone treatments using the hairy-root-inducing bacterium, Agrobacterium rhizogenesandyoungseedlings.TransgenichairyrootsofP.japonicumwereobtainedfromcotyledons2to3weeksafterA. rhizogenes inoculation. The presence and the expression of transgenes in P. japonicum were verified by genomic PCR, Southern blot and RT-PCR methods. Transgenic roots derived from A. rhizogenes-mediated transformation were able to develophaustoriaonriceandmaizeroots.Transgenicrootsalsoformedapparentlycompetenthaustoriainresponseto2,6- dimethoxy-1,4-benzoquinone(DMBQ),ahaustorium-inducingchemical.Usingthissystem,weintroducedareportergene withaCyclinB1promoterintoP.japonicum,andvisualizedcelldivisionduringhaustoriumformation. Conclusions: We provide an easy and efficient method for hairy-root transformation of P. japonicum. Transgenic marker analysis revealed that cell divisions during haustorium development occur 24h after DMBQ treatment. The protocols describedherewillallowfunctionalanalysisofgenesinvolvedinplantparasitism. Citation: Ishida JK, YoshidaS, Ito M, Namba S, Shirasu K (2011) Agrobacteriu