A thiamine-regulatable epitope-tagged protein expression system in fission yeast.
Schizosaccharomyces pombe, the fission yeast, has been a popular and useful model system for investigating the mechanisms of biological processes for a long time. To facilitate purification, localization, and functional analysis of gene products, a wide range of expression vectors have been developed. Several of these vectors utilize the inducible/repressible promoter systems and enable the episomal expression of proteins as fusion proteins with epitope tags attached to their N terminus or C terminus.This chapter provides a detailed protocol for expression of the epitope-tagged proteins from thiamine-regulatable nmt promoter in fission yeast. The yeast culture conditions and procedures for yeast transformation, expression induction, preparation of whole-cell extracts, and analysis of epitope-tagged protein expression by Western blotting are described.
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