Additional vectors for PCR-based gene tagging in Saccharomyces cerevisiae and Schizosaccharomyces pombe using nourseothricin resistance.
The one-step PCR-mediated technique used for modification of chromosomal loci is a powerful tool for functional analysis in yeast. Both Saccharomyces cerevisiae and Schizosaccharomyces pombe are amenable to this technique. However, the scarce availability of selectable markers for Sz. pombe hampers the easy use of this technique in this species. Here, we describe the construction of new vectors deriving from the pFA6a family, which are suitable for tagging in both yeasts owing to the presence of a nourseothricin-resistance cassette. These plasmids allow various gene manipulations at chromosomal loci, viz. N- and C-terminal tagging with 3HA (haemagglutinin) or 13Myc epitopes, GST (glutathione S-transferase), 4TAP (tandem affinity purification) and several GFP (green fluorescent protein) isoforms. For N-terminal modifications, the use of different promoters allows constitutive (PADH1) or regulatable (PGAL1) promoters for S. cerevisiae and derivatives of Pnmt1 for Sz. pombe expression.
Van Driessche
Benoit
B
Unité de Recherche en Biologie Moléculaire, University of Namur, 61 Rue de Bruxelles, 5000 Namur, Belgium.
Tafforeau
Lionel
L
Hentges
Pierre
P
Carr
Antony M
AM
Vandenhaute
Jean
J
D000935:Antifungal Agents / Q000494:pharmacology; D025141:Drug Resistance, Fungal / Q000235:genetics*; D000939:Epitopes / Q000235:genetics; D018390:Gene Targeting; D005819:Genetic Markers; D005822:Genetic Vectors; D010957:Plasmids; D016133:Polymerase Chain Reaction / Q000379:methods*; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000235:genetics*; D012568:Schizosaccharomyces / Q000187:drug effects / Q000235:genetics*; D013309:Streptothricins / Q000494:pharmacology; D014170:Transformation, Genetic
D000935:Antifungal Agents; D000939:Epitopes; D005819:Genetic Markers; D013309:Streptothricins