| Summary: | The use of broad-spectrum antimycotic therapy, immunosuppressive therapy, and indwelling medical devices has contributed to the increased frequency of mucosal and systemic infections caused by <i>Candida glabrata</i>. A major concern for <i>C. glabrata</i> and other <i>Candida</i> spp. infections is the increase in drug resistance. To address these issues, additional molecular tools for the study of <i>C. glabrata</i> are needed. In this investigation, we developed an <i>Agrobacterium tumefaciens</i> transformation system for <i>C. glabrata</i>. A number of parameters were investigated to determine their effect on transformation frequency, and then an optimized protocol was developed. The optimal conditions for the transformation of <i>C. glabrata</i> were found to be an infection incubation temperature of 26 °C, 0.2 mM acetosyringone in both induction media and co-culture media, 0.7% agar concentration, and a multiplicity of infection of 50:1 <i>A. tumefaciens</i> to <i>C. glabrata</i>. Importantly, the frequency of multiple integrations was low (5%), demonstrating that <i>A. tumefaciens</i> generally integrates at single sites in <i>C. glabrata</i>, which is consistent with other fungal <i>A. tumefaciens</i> transformation systems. The development of this system in <i>C. glabrata</i> adds another tool for the molecular manipulation of this increasingly important fungal pathogen.
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