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...

Full description

Saved in:
Bibliographic Details
Published in:Yeast (Chichester, England) England), 2005-10, Vol.22 (13), p.1061-1068
Main Authors: Van Driessche, Benoit, Tafforeau, Lionel, Hentges, Pierre, Carr, Antony M., Vandenhaute, Jean
Format: Article
Language:English
Subjects:
Antifungal Agents - pharmacology
Drug Resistance, Fungal - genetics
epitope tagging
Epitopes - genetics
gene overexpression
Gene Targeting
Genetic Markers
Genetic Vectors
nourseothricin
Plasmids
Polymerase Chain Reaction - methods
Saccharomyces cerevisiae
Saccharomyces cerevisiae - drug effects
Saccharomyces cerevisiae - genetics
Schizosaccharomyces - drug effects
Schizosaccharomyces - genetics
Schizosaccharomyces pombe
Streptothricins - pharmacology
Transformation, Genetic
vector
yeast
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary: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. Copyright © 2005 John Wiley & Sons, Ltd.
ISSN:0749-503X
1097-0061
DOI:10.1002/yea.1293