1. TheAshbya gossypiiEF-1αpromoter of the ubiquitously used MX cassettes is toxic toSaccharomyces cerevisiae
- Author
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Stefan Hohmann, Soode Moghadas Jafari, Marcus Krantz, Martin Zackrisson, Jonas Warringer, Roja Babazadeh, and Anders Blomberg
- Subjects
Genetic Markers ,Green Fluorescent Proteins ,Saccharomyces cerevisiae ,Biophysics ,Heterologous ,Peptide ,Microbial Sensitivity Tests ,Eremothecium ,Biochemistry ,Green fluorescent protein ,Peptide Elongation Factor 1 ,Structural Biology ,Genetics ,Histidine ,Promoter Regions, Genetic ,Molecular Biology ,Gene ,chemistry.chemical_classification ,biology ,MX toxicity ,TAP-tag toxicity ,Cell Biology ,biology.organism_classification ,Molecular biology ,Global strain ,Mutagenesis, Insertional ,chemistry ,Histidine toxicity ,Toxicity ,Protein overexpression ,HIS3 ,Genome, Fungal ,Genetic Engineering ,Plasmids - Abstract
Protein overexpression based on introduction of multiple gene copies is well established. To improve purification or quantification, proteins are typically fused to peptide tags. In Saccharomyces cerevisiae, this has been hampered by multicopy toxicity of the TAP and GFP cassettes used in the global strain collections. Here, we show that this effect is due to the EF-1α promoter in the HIS3MX marker cassette rather than the tags per se. This promoter is frequently used in heterologous marker cassettes, including HIS3MX, KanMX, NatMX, PatMX and HphMX. Toxicity could be eliminated by promoter replacement or exclusion of the marker cassette. To our knowledge, this is the first report of toxicity caused by introduction of a heterologous promoter alone.
- Published
- 2011
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