Seamless Gene Tagging by Endonuclease-Driven Homologous Recombination 英文参考文献.docVIP

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Seamless Gene Tagging by Endonuclease-Driven Homologous Recombination 英文参考文献.doc

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Seamless Gene Tagging by Endonuclease-Driven Homologous Recombination 英文参考文献

SeamlessGeneTaggingbyEndonuclease-Driven HomologousRecombination AntonKhmelinskii1.¤,MatthiasMeurer1.¤,NurlanbekDuishoev1,NicolasDelhomme2,MichaelKnop1*¤ 1CellBiologyandBiophysicsUnit,EuropeanMolecularBiologyLaboratory(EMBL),Heidelberg,Germany,2GenomeBiologyUnit,EuropeanMolecularBiologyLaboratory (EMBL),Heidelberg,Germany Abstract Gene tagging facilitates systematic genomic and proteomic analyses but chromosomal tagging typically disrupts gene regulatorysequences.Herewedescribeaseamlessgenetaggingapproachthatpreservesendogenousgeneregulationand ispotentiallyapplicableinanyspecieswithefficientDNAdouble-strandbreakrepairbyhomologousrecombination.We implement seamless tagging in Saccharomyces cerevisiae and demonstrate its application for protein tagging while preservingsimultaneouslyupstreamanddownstreamgeneregulatoryelements.Seamlesstaggingiscompatiblewithhigh- throughputstrainconstructionusingsyntheticgeneticarrays(SGA),enablesfunctionalanalysisoftranscriptionantisenseto openreadingframesandshouldfacilitatesystematicandminimally-invasiveanalysisofgenefunctions. Citation:KhmelinskiiA,MeurerM,DuishoevN,DelhommeN,KnopM(2011)SeamlessGeneTaggingbyEndonuclease-DrivenHomologousRecombination.PLoS ONE6(8):e23794.doi:10.1371/journal.pone.0023794 Editor:KevinG.Hardwick,UniversityofEdinburgh,UnitedKingdom ReceivedJune24,2011;AcceptedJuly27,2011;PublishedAugust22,2011 Copyright: ?2011Khmelinskiietal.Thisisanopen-access articledistributedunderthetermsoftheCreativeCommonsAttributionLicense,whichpermits unrestricteduse,distribution,andreproductioninanymedium,providedtheoriginalauthorandsourcearecredited. Funding:ThisworkwassupportedbyaLong-TermfellowshipfromtheEuropeanMolecularBiologyOrganization(EMBOALTF1124-2010)toA.K.Thefunders hadnoroleinstudydesign,datacollectionandanalysis,decisiontopublish,orpreparationofthemanuscript. CompetingInterests:Theauthorshavedeclaredthatnocompetinginterestsexist. *E-mail:m.knop@zmbh.uni-heidelberg.de .Theseauthorscontributedequallytothiswork.