developmental biology protocols - rocky s. tuan and cecilia w. lo

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1、MethodsinMolecularBiologyMethodsinMolecularBiologyTMTMVOLUME137DevelopmentalDevelopmentalBiologyBiologyProtocolsProtocolsVVolumeIIIolumeIIIEditedbyEditedbyRockyS.TuanRockyS.TuanCeciliaW.LoCeciliaW.LoHUMANAPRESSOverview31DevelopmentalBiologyProtocolsOverviewI

2、IIRockyS.TuanandCeciliaW.Lo1.IntroductionThemarriageofcellandmolecularbiologywithembryologyhasproducedremark-ableadvancesforthefieldofdevelopmentalbiology.InthisthirdvolumeofDevelop-mentalBiologyProtocols,contemporary,practicalmethodsarefirstpresentedforthea

3、nalysisandmanipulationofdevelopmentalgeneexpression.Toillustratehowsuchtechniques,aswellasproceduresofexperimentalembryologyincludingthosedescribedinthefirsttwovolumesoftheseries,maybeappliedinthestudyofdevelop-ment,apanoramiccollectionofexperimentalmodelsof

4、morphogenesis,development,andcellulardifferentiationaredetailed.Bothinvivoandinvitrosystemsareincluded.Thevolumeconcludeswithvariousexamplesofdevelopmentalmodelsofdiseasesandtheirmolecularbasis.2.ManipulationofDevelopmentalGeneExpressionandFunctionDrosophila

5、hasbeenandremainsoneofthemostversatilemodelsystemsforthemanipulationofdevelopmentalgeneexpression.Chapter2focusesonadescriptionoftheexperimentalapproachescurrentlyusedinectopicgeneexpressioninDrosophilatoexaminethefunctionofagivengeneinthedesiredtissue.Chapt

6、er3dealswiththeutilizationofthehighlyefficientFLP/FRTyeastsite-specificrecombinationsystemtogeneratesomaticandgermlineclonesinDrosophilaforphenotypicanalysisandscreening.Chapters3and4addressthemethodsusedtoaltergeneexpressionaswellasgenefunctioninanotherexpe

7、rimentallyhighlyaccessiblesystem,thedevelopingchickembryo.Chapter3describestheapplicationofantisenseoligonucleotidesto“knockdown”geneexpressioninsomiticstagechickembryos,whereasChapter4discusseshowfunctionalneutralizingmonoclonalantibodiesmaybeusedtoblockthe

8、activityofaspecificgeneproduct,N-cadherin,inthedevelopingchickembryoniclimbbud.3.AnalysisofGeneExpressionThefirststepinanalyzingthemolecularbasisofanydevelopmentaleventistocharacterizeandcompare