TABLE 1.
Strains and plasmids used in this study
| Strain or plasmid | Description |
|---|---|
| Strains | |
| MH2-d | MATa/α ade2-1/ade2-1 his3Δ200/his3Δ200 leu2-1/leu2-1 ura3-52/ura3-52 trp1Δ101/trp1Δ101 gal2/gal2 |
| MH2-h | MATα ade2-1 his3,200 leu2-1 ura3-52 trp1,101 gal2 |
| YTK84 | MH2-d rvb2::HIS3/RVB2 |
| YTK86 | MH2-h rvb2::HIS3 pTK123 (RVB2 URA3 2 μm) |
| YWD312 | YTK86 Rvb2-GFP (cured of pTK123) |
| YWD313 | YTK86 pGAL-rvb2 (TRP1 CEN6) (cured of pTK123) |
| YWD339 | MH2-h pYES235W (URA3 2 μm) |
| YWD340 | MH2-h pWD296N (URA3 2μm) |
| YTK100 | YTK86 pTK149 (TRP1 CEN6) (cured of pTK123) |
| YTK104 | YTK86 pTK153 (TRP1 CEN6) (cured of pTK123) |
| Plasmids | |
| pTK123 | PCR product of RVB2 with natural flanking sequence inserted into the BamHI site of Yep24 (URA3 2μm) |
| pWD235WGFP | Yeast RVB2 ORF fused in frame to GFP (TRP1 CEN6) |
| GFP/pCDNA3.1 | Super GFP ORF under control of CMV promoter (Invitrogen) |
| pTK135 | RVB2 gene with natural flanking sequences inserted into BamHI site of pRS314 (TRP1 CEN6) |
| pTK147 | PCR-amplified mouse Rvb2 homolog (p50) ORF inserted into MCS of pcDNA 3.1/CT-GFP TOPO (Invitrogen) |
| pTK149 | pTK135 with rvb2 K81A mutation |
| pTK151 | pTK135 with rvb2 G75A mutation |
| pTK153 | pTK135 with rvb2 G80A mutation |
| pTK156 | pTK135 with rvb2 K81R mutation |
| pTK157 | pTK135 with rvb2 T82S mutation |
| pTK159 | pTK135 with rvb2 I295V mutation |
| pTK163 | pTK135 with rvb2 E297D mutation |
| pTK165 | pTK135 with rvb2 V298G mutation |
| pRSY14 | pRS314 with GAL1 promoter and CYC-1 terminator |
| pYES235W | pYES2 with wild-type RVB2 gene |
| pWD296N | Plasmid pYES235W with rvb2 D296N mutation |
| pGAL-rvb2 | RVB2 ORF flanked by GAL1 promoter and CYCI terminator (TRP1 CEN6) |
| pYDp-H | Plasmid carrying HIS3 gene and regulatory sequences (Amp; ori); PCR template for gene replacement |
| Rvb2-GFP | Yeast Rvb2 fused in frame to the N terminus of GFP (TRP1/CEN6) |