New pFA‐cassettes for PCR‐based gene manipulation in Candida albicans

Several modules for efficient PCR‐based gene disruption have recently been introduced in Candida albicans . These are based on auxotrophic marker genes for deficient strains derived from SC5314/CAI4. Commonly used protocols for the transformation C. albicans are based either on the lithium acetate procedure or on electroporation also used for Saccharomyces cerevisiae . Here we present our updated arsenal of pFA‐modules that now include the heterologous marker genes HIS1 from C. dubliniensis and LEU2 from C. maltosa ( Noble and Johnson 2005) and the dominant selection marker ca SAT1 (Reuss et al. 2004). We also introduce the Ashbya gossypii TEF1 ‐promoter as a strong constitutive promoter. With these new elements an enlarged collection of pFA‐marker and pFA‐marker‐promoter modules were generated containing 17 new modules. In addition, N‐terminal tagging with GFP‐(GA) 6 and epitope‐tagging modules using the 6 ×‐HIS‐tag were constructed. This adds to the previous modules that only enabled C‐terminal GFP‐tagging of genes ( Gola et al. 2003). In total 29 pFA‐modules are currently freely available from our lab which – together with an update on the diagnostic verification procedure – further enlarge the C. albicans molecular toolbox and enhance our capabilities to use PCR‐based gene alteration methods in C. albicans . (© 2006 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)