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Additional vectors for PCR‐based gene tagging in Saccharomyces cerevisiae and Schizosaccharomyces pombe using nourseothricin resistance
Author(s) -
Van Driessche Benoit,
Tafforeau Lionel,
Hentges Pierre,
Carr Antony M.,
Vandenhaute Jean
Publication year - 2005
Publication title -
yeast
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.923
H-Index - 102
eISSN - 1097-0061
pISSN - 0749-503X
DOI - 10.1002/yea.1293
Subject(s) - biology , gene , botany , microbiology and biotechnology , genetics
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 ( h aem a gglutinin) or 13Myc epitopes, GST ( g lutathione S ‐ t ransferase), 4TAP ( t andem a ffinity p urification) and several GFP ( g reen f luorescent p rotein) isoforms. For N‐terminal modifications, the use of different promoters allows constitutive ( P ADH 1 ) or regulatable ( P GAL 1 ) promoters for S. cerevisiae and derivatives of P nmt 1 for Sz. pombe expression. Copyright © 2005 John Wiley & Sons, Ltd.