Figure 1

The Degron cassette enables the construction of potential td alleles for any gene. The cassette contains KanMX for selection (G418 resistance), the CUP1 promoter (P_CUP1) that is expressed in the presence of copper and repressed in its absence, and the gene encoding the temperature-sensitive Degron (td) protein fused to the HA epitope. A gene-specific pair of oligonucleotides (A and B), with 3' ends matching the cassette (shown by vertical dashed lines), is used to amplify the cassette by PCR. Because the 5' end of each oligonucleotide matches sequences of gene X (shown by slanted dashed lines), transformation of the cassette into yeast cells targets the cassette for insertion upstream of the coding sequence by homologous recombination, replacing the native promoter and creating a gene X-td fusion. Materials and details of construction are available at [20].