TABLE 2

Plasmids used in this study

PlasmidGenotypeDescription/reference
pEGKGGAL1prom-GST 2μm URA3162
pEGKG-Rod1GAL1prom-GST 2μm URA3163
pEGKG-Rog3GAL1prom-GST 2μm URA3163
pEGKG-Ldb19GAL1prom-GST 2μm URA3163
pEGKG-Art5GAL1prom-GST 2μm URA3163
pEGKG-Rod1PANA (pJT4954)aGAL1prom-GST 2μm URA3This study
pEGKG-Rod1PASA (pJT4955)aGAL1prom-GST 2μm URA3This study
pEGKG-Rod1PPXY-less (pJT4956)aGAL1prom-GST 2μm URA3This study
pEGKG-Rod14KR (Rod1K235R K245R K264R K267R) (pJT5045)aGAL1prom-GST 2μm URA3This study
pEGKG-Rog3PANA (pJT4958)aGAL1prom-GST 2μm URA3This study
pEGKG-Rog3PASA (pJT4959)aGAL1prom-GST 2μm URA3This study
pEGKG-Rog3PPXY-less (pJT4960)aGAL1prom-GST 2μm URA3This study
pEGKG-Rog3VASA (pJT4978)aGAL1prom-GST 2μm URA3This study
pEGKG-Rog3V/PPXY-less (pJT4979)aGAL1prom-GST 2μm URA3This study
pEGKG-Rog3Δ400 (pJT4983)aGAL1prom-GST 2μm URA3This study
pEGKG-Rog34KR (Rog3K235R K245R K264R K267R) (pJT5060)aGAL1prom-GST 2μm URA3This study
pEGKG-Rod1AQAKAA (pJT4957)aGAL1prom-GST 2μm URA3This study
pRS313CEN HIS3164
pRS313-Ldb19 (pJT4963)bLDB19prom CEN HIS3This study
pRS313-Ldb19PAIA (pJT4964)cLDB19prom CEN HIS3This study
pRS313-Ldb19 PPCY (pJT4965)cLDB19prom CEN HIS3This study
pRS313-Ldb19PPXY-less (pJT4966)cLDB19prom CEN HIS3This study
pRS313-Ldb19K486R (pJT5001)cLDB19prom CEN HIS3This study
pGEX4T1-GST-Ste2297–431T7 AMP163
  • a Generated by site-directed mutagenesis (67) with synthetic oligonucleotides containing the desired codon alterations (using the wild-type sequence in pRS313 vectors as the template). DNA from the corresponding gene was amplified from genomic DNA by PCR (66) and then cloned into pEGKG.

  • b DNA of the corresponding gene was amplified from genomic DNA by PCR (66) and then cloned into the XmaI-NotI sites in pRS313 (164).

  • c Generated by site-directed mutagenesis with synthetic oligonucleotides containing the desired codon alterations and the corresponding LDB19 DNA inserted into pRS313 as the template.