Abstract
The conditional control of protein expression is useful to characterize the function of proteins, especially of those that are essential for cell viability. Two degron-based systems, temperature-sensitive and auxin-inducible degrons, can be used to generate conditional mutants of budding yeast, simply by transforming appropriate cells with PCR-amplified DNA. We describe a protocol for the generation of temperature-sensitive and auxin-inducible degron mutants. We also show that a conditional mutant with few spontaneous revertants was generated by combining two degron systems for the Inn1 protein. Finally, we describe a suppressor screening method that uses the dual degron-Inn1 mutant to identify mutant proteins that suppress Inn1-K31A, which has a defect in cytokinesis.
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