Yeast Genetic Transformation Protocols

Wild-type S. cerevisiae and wild-type C. glabrata were used as the host strains for GRC transformations as both are ura3⁻his3⁻ (See Table 1 for strains). Deletions of RAD52 and DNL4 in both S. cerevisiae and C. glabrata were generated using antibiotic resistance genes KANMX6 and NATMX6 (conferring resistance to G-418 and nourseothricin, respectively) and homologous recombination to delete the ORFs [5] (link), [15] (link), [16] (link), which was confirmed by PCR. For transformations, yeast strains were grown in YEPD medium at 30°C until logarithmic growth phase (OD₆₀₀ 0.2–0.5). To select for plasmids, strains were grown in synthetic dextrose (SD) medium with CSM lacking the appropriate amino acids (either histidine or uracil) (Sunrise Science, San Diego, CA, USA). Transformations were performed using a standard lithium acetate protocol [17] (link), [18] , [19] (link).