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Gene Activation Using FLP Recombinase in C. elegans 英文参考文献
GeneActivationUsingFLPRecombinaseinC.elegans
M.WayneDavis1,2,J.JasonMorton3¤,DanaCarroll3,ErikM.Jorgensen1,2*
1HowardHughesMedicalInstitute,UniversityofUtah,SaltLakeCity,Utah,UnitedStatesofAmerica,2DepartmentofBiology,UniversityofUtah,SaltLakeCity,Utah,
UnitedStatesofAmerica,3DepartmentofBiochemistry,UniversityofUtahSchoolofMedicine,SaltLakeCity,Utah,UnitedStatesofAmerica
Abstract
TheFLPenzymecatalyzesrecombinationbetweenspecifictargetsequencesinDNA.HereweuseFLPtotemporallyand
spatiallycontrolgeneexpressioninthenematodeC.elegans.Transcriptionisblockedbythepresenceofan‘‘offcassette’’
betweenthepromoterandthecodingregionofthedesiredproduct.The‘‘offcassette’’iscomposedofatranscriptional
terminatorflankedbyFLPrecognitiontargets(FRT).ThissequencecanbeexcisedbyFLPrecombinasetobringtogetherthe
promoterandthecodingregion.Wehaveintroducedtwofluorescentreportersintothesystem:aredreporterforpromoter
activity prior to FLP expression and a green reporter for expression of the gene of interest after FLP expression. The
constructsaredesignedusingthemultisiteGatewaysystem,sothatpromotersandcodingregionscanbequicklymixed
and matched. We demonstrate that heat-shock-driven FLP recombinase adds temporal control on top of tissue specific
expression provided by the transgene promoter. In addition, the temporal switch is permanent, rather than acute, as is
usually the case for heat-shock driven transgenes. Finally,FLP expressioncan be driven by a tissue specific promoter to
provideexpressioninasubsetofcellsthatcanonlybeaddressedastheintersectionoftwoavailablepromoters.Asatestof
the system, we have driven the light chain of tetanus toxin, a protease that cleaves the synaptic vesicle protein
synaptobrevin.Weshowthatwecanusethistoinactivatesynaptictransmissioninallneuronsorasubsetofneuronsina
FLP-dependentmanner.
Citation:DavisMW,MortonJJ,CarrollD,JorgensenEM(2008)GeneActivationUsingFLPRecombinaseinC.elegans.PLoSGenet4(3):e1000028.doi:10.1371/
journal.pgen.1000028
Editor:StuartK.Kim,Stanfo
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