Abstract
Expression plasmids for Saccharomyces cerevisiae offer a wide choice of vector copy number, promoters of varying strength and selection markers. These expression plasmids are usually shuttle vectors that can be propagated both in yeast and bacteria, making them useful in gene cloning. For heterologous production of membrane proteins, we used the green fluorescent protein (GFP) fusion technology which was previously developed in the Escherichia coli system. We designed an expression plasmid carrying an inducible GAL1 promoter, a gene encoding a membrane protein of interest and the GFP-octa-histidine sequence. Here we describe construction of multi-copy yeast expression plasmids by homologous recombination in S. cerevisiae.
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Abbreviations
- GAL:
-
Galactokinase
- GPD:
-
Glyceraldehyde-3-phosphate dehydrogenase
- TEF:
-
Translation elongation factor 1α
- TEV site:
-
Tobacco etch virus protease cleavage site
- yEGFP:
-
Yeast-enhanced green fluorescent protein
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Acknowledgments
This work was supported by the Royal Society (United Kingdom) through a University Research Fellowship to DD and by a Basic Science Research Program grant through the National Research Foundation of Korea (NRF) funded by the Ministry of Education, Science and Technology (NRF0409-20100093) to HK.
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Drew, D., Kim, H. (2012). Preparation of Saccharomyces cerevisiae Expression Plasmids. In: Bill, R. (eds) Recombinant Protein Production in Yeast. Methods in Molecular Biology, vol 866. Humana Press. https://doi.org/10.1007/978-1-61779-770-5_4
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DOI: https://doi.org/10.1007/978-1-61779-770-5_4
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