TABLE 1.
Yeast strains and plasmids used in this study
| Yeast strain or plasmid | Genotype or description | Source or reference |
|---|---|---|
| Strains | ||
| MHY501 | MATα his3-Δ200 leu2-3, 112 trp1-1 ura3-52 lys2-801 gal2 | M. Hochstrasser |
| YPH499 | MATaade2-101 his3-Δ200 leu2-Δ1 trp1-Δ63 ura3-52 lys2-801 | Stratagene |
| 15DaubΔ | MATabar1Δ ade1 his2 leu2-3, 112 trp1-1 ura3Δns | D. Stuart |
| DSY1105 | MATabar1Δ ade1 his2 leu2-3, 112 trp1-1 ura3Δns cdc53-1 | D. Stuart |
| Plasmids | ||
| Wild-type and control plasmids | ||
| pJD325 | 2μm PCUP1-UBA1-6xHis [LEU2] | D. Findley |
| pES5 | 2μm PCUP1-HA-Ub [URA3] | This study |
| pESC200 | 2μm PGAL1-CDC53-3xHA [URA3] | This study |
| pESC(Trp) | 2μm PGAL1-Myc, PGAL10-Flag [TRP1] | Stratagene |
| pESC(Ura) | 2μm PGAL1-Myc, PGAL10-Flag [URA3] | Stratagene |
| pESC1 | 2μm PGAL10-Flag-CDC34 [TRP1] | This study |
| pESC2 | 2μm PGAL1-Myc-CDC34 [TRP1] | This study |
| pESCA | 2μm PGAL1-Myc-CDC34, PGAL10-Flag-CDC34 [TRP1] | This study |
| Cdc34 catalytic domain derivative plasmids | ||
| pESC15 | 2μm PGAL10-Flag-cdc34 C95A [TRP1] | This study |
| pESC16 | 2μm PGAL1-Myc-cdc34 C95A [TRP1] | This study |
| pESCH | 2μm PGAL1-Myc-cdc34 C95A, PGAL10-Flag-cdc34 C95A [TRP1] | This study |
| pESC19 | 2μm PGAL10-Flag-cdc34 S97D [TRP1] | This study |
| pESC20 | 2μm PGAL1-Myc-cdc34 S97D [TRP1] | This study |
| pESCJ | 2μm PGAL1-Myc-cdc34 S97D, PGAL10-Flag-cdc34 S97D [TRP1] | This study |
| pESC3 | 2μm PGAL10-Flag-cdc34 S73K/S97D [TRP1] | This study |
| pESC4 | 2μm PGAL1-Myc-cdc34-S73K/S97D [TRP1] | This study |
| pESCB | 2μm PGAL1-Myc-cdc34 S73K/S97D, PGAL10-Flag-cdc34 S73K/S97D [TRP1] | This study |
| pESC5 | 2μm PGAL10-Flag-cdc34Δ12 [TRP1] | This study |
| pESC6 | 2μm PGAL1-Myc-cdc34Δ12 [TRP1] | This study |
| pESCC | 2μm PGAL1-Myc-cdc34Δ12, PGAL10-Flag-cdc34 Δ12 [TRP1] | This study |
| pESC7 | 2μm PGAL10-Flag-cdc34 S73K/S97D/Δ 12 [TRP1] | This study |
| pESC8 | 2μm PGAL1-Myc-cdc34 S73K/S97D/Δ 12 [TRP1] | This study |
| pESCD | 2μm PGAL1-Myc-cdc34 S73K/S97D/Δ 12, PGAL10-Flag-cdc34 S73K/S97D/Δ 12 [TRP1] | This study |
| Cdc34 carboxy-terminal truncation plasmids | ||
| pESC9 | 2μm PGAL10-Flag-cdc34Δ 170[TRP1] | This study |
| pESC10 | 2μm PGAL1-Myc-cdc34Δ 170[TRP1] | This study |
| pESCE | 2μm PGAL1-Myc-cdc34Δ 170, PGAL10-Flag-cdc34Δ 170 [TRP1] | This study |
| pESC11 | 2μm PGAL10-Flag-cdc34Δ 185[TRP1] | This study |
| pESC12 | 2μm PGAL1-Myc-cdc34Δ 185[TRP1] | This study |
| pESCF | 2μm PGAL1-Myc-cdc34Δ 185, PGAL10-Flag-cdc34 Δ185 [TRP1] | This study |
| pESC13 | 2μm PGAL10-Flag-cdc34Δ 209[TRP1] | This study |
| pESC14 | 2μm PGAL1-Myc-cdc34Δ 209[TRP1] | This study |
| pESCG | 2μm PGAL1-Myc-cdc34Δ 209, PGAL10-Flag-cdc34 Δ209 [TRP1] | This study |
| Bacterial expression plasmids | ||
| pREGB1 | pET-3a derivative + Ub | 15 |
| pREGB6 | pET-3a derivative + CDC34 | 26 |
| pCdc34Δ185 | pET-3a derivative + cdc34Δ185 | 26 |
| pCdc34Δ209 | pET-3a derivative + cdc34Δ209 | 26 |
| pBV4 | pET-3a derivative + cdc34C95A | This study |
| pBV8 | pET-3a derivative + cdc34S97D | This study |
| pBV9 | pET-3a derivative + cdc34S73K/S97D | This study |