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  • Research Paper
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A System for Dominant Transformation and Plasmid Amplification in Saccharomyces cerevisiae

Abstract

The cDNA of the mouse dihydrofolate reductase (dhfr) was put under transcriptional control of the yeast cytochrome c or the actin promoter and cytochrome c terminator on 2μ-derived plasmids. Transformation of yeast cells with such plasmids made them resistant to the antime-tabolite methotrexate. Yeast cells were transformed by direct drug-resistant selection and the plasmids amplified in both enriched and defined media. The transformation efficiency of the recombinant plasmids containing the mouse dhfr gene by direct methotrexate selection was similar to the efficiency obtained by auxotrophic selection. The copy number amplification was 3 to 4-fold. The system was used to study the expression of the human interleukin 2 gene, placed behind the triose phosphate isomerase promoter. This allowed synthesis of mature human interleukin 2 at a level of about 5 percent of total protein in defined medium.

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Zhu, J., Contreras, R., Gheysen, D. et al. A System for Dominant Transformation and Plasmid Amplification in Saccharomyces cerevisiae. Nat Biotechnol 3, 451–456 (1985). https://doi.org/10.1038/nbt0585-451

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