Zeocin resistance as a dominant selective marker for transformation and targeted gene deletions in Candida glabrata

Many of the genetic tools used to generate gene knockouts in Candida glabrata exploit auxotrophic markers but this is not suitable for use with clinical strains. Antibiotic resistance markers, however, allow one to target genes to be deleted without any prior genetic manipulation of clinical isolates. Such antibiotic selection markers have been widely reported for the manipulation of Saccharomyces cerevisiae. However, very few antibiotic resistance markers have been shown to be useful in C. glabrata. Here, we report the use of Zeocin resistance (Zeo(R)), encoded by the ble gene from Streptoalloteichus hindustanus, as a new positive selection marker for the genetic manipulation of C. glabrata including clinical strains that we show are significantly more sensitive to Zeocin than to G418. The potential of the Zeo(R) marker for targeted gene disruption in C. glabrata was confirmed by constructing deletions of the ADE2 in both a laboratory and a clinical strain of C. glabrata, using both short (90 bp) and long (400 bp) homology cassettes.