RRC ID 30952
Author Levi S, Polyakov M, Egelhoff TT.
Title Green fluorescent protein and epitope tag fusion vectors for Dictyostelium discoideum.
Journal Plasmid
Abstract We have constructed expression vectors for Dictyostelium discoideum which encode a green fluorescent protein (GFP) sequence upstream of a multicloning site for introduction of sequences of interest. Insertion of cDNAs into the multicloning site results in expression of fusion protein bearing an amino- or carboxyl-terminal GFP tag which can be used for fluorescent localization studies in Dictyostelium cells. A parallel construct fuses a FLAG epitope tag at the amino terminus of expressed protein. Each fusion cartridge was placed either in a G418-resistance vector allowing transactivated Ddp2-based extrachromosomal replication or in a vector allowing autonomous Ddp1-based replication. Distinct differences in expression stability were observed in the two vector types. When GFP-expressing cells were analyzed by fluorescence microscopy, significant cell-to-cell variability in expression level was observed when expression was based on the Ddp2 vector, while less cell-to-cell variation in expression level was observed when the Ddp1 backbone was used for expression.
Volume 44(3)
Pages 231-8
Published 2000-11-1
DOI 10.1006/plas.2000.1487
PII S0147-619X(00)91487-2
PMID 11078649
MeSH Amino Acid Sequence Animals Bacterial Proteins / genetics* Bacterial Proteins / metabolism Base Sequence Cloning, Molecular / methods* Dictyostelium / genetics* Dictyostelium / metabolism Escherichia coli Proteins* Genetic Vectors / genetics* Green Fluorescent Proteins Luminescent Proteins / biosynthesis Luminescent Proteins / genetics* Molecular Sequence Data Oligopeptides Oxidoreductases* Peptides / genetics* Peptides / metabolism Plasmids / genetics* Recombinant Fusion Proteins / biosynthesis Recombinant Fusion Proteins / genetics*
IF 2.805
Times Cited 130
WOS Category MICROBIOLOGY GENETICS & HEREDITY
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