Cnidarian and Bilaterian Promoters Can Direct GFP Expression in Transfected Hydra

Developmental Biology

Brigitte Galliot¹  Marijana Miljkovic¹  Françoise Mazet¹ 

[1] Department of Zoology and Animal Biology, University of Geneva, 30 Quai Ernest Ansermet, CH-1211, Genève 4, Switzerland

关键词: cnidaria;    hydra;    electroporation;    transfection;    GFP;    ectoderm;    endoderm;    cnox-2 promoter;    actin promoter;    ribosomal gene promoters;   

DOI  :  10.1006/dbio.2002.0676

学科分类：生物科学（综合）

来源: Academic Press

【 摘 要 】

Completesexualdevelopmentisnoteasilyamenabletoexperimentationinhydra.Therefore,theanalysisofgenefunctionandgeneregulationrequirestheintroductionofexogenousDNAinalargenumberofcellsofthehydrapolypsandthesignificantexpressionofreporterconstructsinthesecells.WepresentheretheprocedurewherebywecoupledDNAinjectionintothegastriccavitytoelectroporationofthewholeanimalinordertoefficientlytransfecthydrapolyps.WecoulddetectGFPfluorescenceinbothendodermalandectodermalcelllayersofliveanimalsandinepithelialaswellasinterstitialcelltypesofdissociatedhydra.Inaddition,wecouldconfirmGFPproteinexpressionbyshowingcolocalisationbetweenGFPfluorescenceandanti-GFPimmunofluorescence.Finally,whenaFLAGepitopewasinsertedin-framewiththeGFPcodingsequence,GFPfluorescencealsocolocalisedwithanti-FLAGimmunofluorescence.ThisGFPexpressioninhydracellswasdirectedbyvariouspromoters,eitherhomologous,likethehydrahomeoboxcnox-2genepromoter,orheterologous,likethetwonematoderibosomalproteinS5andL28genepromoters,andthechickenβ-actingenepromoter.Thisstrategyprovidesnewtoolsfordissectingdevelopmentalmolecularmechanismsinhydra;morespecifically,thegeneticregulationsthattakeplaceinendodermalcellsatthetimebuddingorregenerationisinitiated.

【 授权许可】

Unknown