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Characterization of different promoters for designing a new expression vector in Saccharomyces cerevisiae
Author(s) -
Partow Siavash,
Siewers Verena,
Bjørn Sara,
Nielsen Jens,
Maury Jérôme
Publication year - 2010
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.1806
Subject(s) - promoter , saccharomyces cerevisiae , lac operon , biology , expression vector , reporter gene , plasmid , galactose , gene , shuttle vector , gene expression , beta galactosidase , microbiology and biotechnology , vector (molecular biology) , genetics , biochemistry , recombinant dna
The widely used pESC vector series (Stratagene, La Jolla, CA, USA) with the bidirectional GAL1 / GAL10 promoter provides the possibility of simultaneously expressing two different genes from a single vector in Saccharomyces cerevisiae . This system can be induced by galactose and is repressed by glucose. Since S. cerevisiae prefers glucose as a carbon source, and since its growth rate is higher in glucose than in galactose‐containing media, we compared and evaluated seven different promoters expressed during growth on glucose (p TEF1 , p ADH1 , p TPI1 , p HXT7 , p TDH3 , p PGK1 and p PYK1 ) with two strong galactose‐induced promoters (p GAL1 and p GAL10 ), using lacZ as a reporter gene and measuring LacZ activity in batch and continuous cultivation. TEF1 and PGK1 promoters showed the most constant activity pattern at different glucose concentrations. Based on these results, we designed and constructed two new expression vectors which contain the two constitutive promoters, TEF1 and PGK1 , in opposite orientation to each other. These new vectors retain all the features from the pESC–URA plasmid except that gene expression is mediated by constitutive promoters. Copyright © 2010 John Wiley & Sons, Ltd.