The Ashbya gossypii EF-1α promoter of the ubiquitously used MX cassettes is toxic to Saccharomyces cerevisiae.
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.
Department of Cell and Molecular Biology, University of Gothenburg, Gothenburg, Sweden.
Roja
R
Babazadeh
Soode Moghadas
SM
Jafari
Martin
M
Zackrisson
Anders
A
Blomberg
Stefan
S
Hohmann
Jonas
J
Warringer
Marcus
M
Krantz
D055169:Eremothecium / Q000201:enzymology / Q000235:genetics*; D005818:Genetic Engineering / Q000009:adverse effects / Q000379:methods*; D005819:Genetic Markers / Q000235:genetics; D016681:Genome, Fungal / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000235:genetics; D006639:Histidine / Q000096:biosynthesis; D008826:Microbial Sensitivity Tests; D016254:Mutagenesis, Insertional / Q000235:genetics*; D020648:Peptide Elongation Factor 1 / Q000235:genetics*; D010957:Plasmids / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000235:genetics*; D012441:Saccharomyces cerevisiae / Q000166:cytology* / Q000235:genetics*
D005819:Genetic Markers; D020648:Peptide Elongation Factor 1; D049452:Green Fluorescent Proteins; D006639:Histidine