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dc.contributor.authorJanke, Carsten
dc.contributor.authorMagiera, Maria M
dc.contributor.authorRathfelder, Nicole
dc.contributor.authorTaxis, Christof
dc.contributor.authorReber, Simone
dc.contributor.authorMaekawa, Hiromi
dc.contributor.authorMoreno-Borchart, Alexandra
dc.contributor.authorDoenges, Georg
dc.contributor.authorSchwob, Etienne
dc.contributor.authorSchiebel, Elmar
dc.contributor.authorKnop, Michael
dc.date.accessioned2009-08-25T11:16:17Z
dc.date.available2009-08-25T11:16:17Z
dc.date.issued2004-08
dc.identifier.citationA versatile toolbox for PCR-based tagging of yeast genes: new fluorescent proteins, more markers and promoter substitution cassettes. 2004, 21 (11):947-62 Yeasten
dc.identifier.issn0749-503X
dc.identifier.pmid15334558
dc.identifier.doi10.1002/yea.1142
dc.identifier.urihttp://hdl.handle.net/10541/78447
dc.description.abstractTagging of genes by chromosomal integration of PCR amplified cassettes is a widely used and fast method to label proteins in vivo in the yeast Saccharomyces cerevisiae. This strategy directs the amplified tags to the desired chromosomal loci due to flanking homologous sequences provided by the PCR-primers, thus enabling the selective introduction of any sequence at any place of a gene, e.g. for the generation of C-terminal tagged genes or for the exchange of the promoter and N-terminal tagging of a gene. To make this method most powerful we constructed a series of 76 novel cassettes, containing a broad variety of C-terminal epitope tags as well as nine different promoter substitutions in combination with N-terminal tags. Furthermore, new selection markers have been introduced. The tags include the so far brightest and most yeast-optimized version of the red fluorescent protein, called RedStar2, as well as all other commonly used fluorescent proteins and tags used for the detection and purification of proteins and protein complexes. Using the provided cassettes for N- and C-terminal gene tagging or for deletion of any given gene, a set of only four primers is required, which makes this method very cost-effective and reproducible. This new toolbox should help to speed up the analysis of gene function in yeast, on the level of single genes, as well as in systematic approaches.
dc.language.isoenen
dc.subject.meshGene Targeting
dc.subject.meshGenetic Markers
dc.subject.meshLuminescent Proteins
dc.subject.meshPlasmids
dc.subject.meshPolymerase Chain Reaction
dc.subject.meshPromoter Regions, Genetic
dc.subject.meshRecombination, Genetic
dc.subject.meshRestriction Mapping
dc.subject.meshSaccharomyces cerevisiae
dc.subject.meshSaccharomyces cerevisiae Proteins
dc.subject.meshTransformation, Genetic
dc.titleA versatile toolbox for PCR-based tagging of yeast genes: new fluorescent proteins, more markers and promoter substitution cassettes.en
dc.typeArticleen
dc.contributor.departmentCRBM, CNRS FRE2593, 1919 Route de Mende, F-34293 Montpellier cedex 5, France.en
dc.identifier.journalYeasten
html.description.abstractTagging of genes by chromosomal integration of PCR amplified cassettes is a widely used and fast method to label proteins in vivo in the yeast Saccharomyces cerevisiae. This strategy directs the amplified tags to the desired chromosomal loci due to flanking homologous sequences provided by the PCR-primers, thus enabling the selective introduction of any sequence at any place of a gene, e.g. for the generation of C-terminal tagged genes or for the exchange of the promoter and N-terminal tagging of a gene. To make this method most powerful we constructed a series of 76 novel cassettes, containing a broad variety of C-terminal epitope tags as well as nine different promoter substitutions in combination with N-terminal tags. Furthermore, new selection markers have been introduced. The tags include the so far brightest and most yeast-optimized version of the red fluorescent protein, called RedStar2, as well as all other commonly used fluorescent proteins and tags used for the detection and purification of proteins and protein complexes. Using the provided cassettes for N- and C-terminal gene tagging or for deletion of any given gene, a set of only four primers is required, which makes this method very cost-effective and reproducible. This new toolbox should help to speed up the analysis of gene function in yeast, on the level of single genes, as well as in systematic approaches.


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