title: Development of a Candida glabrata dominant nutritional transformation marker utilizing the Aspergillus nidulans acetamidase gene (amdS).
abstract: The gene encoding Aspergillus nidulans acetamidase (amdS) was placed under control of Candida albicans ACT1 promoter and terminator sequences and then cloned into a plasmid containing C. glabrata ARS10,CEN8 or ARS10+CEN8 sequences. All plasmids transformed C. glabrata wild-type cells to acetamide+, with the ARS-only containing plasmid transforming cells at the highest frequencies (>1.0 × 10(4) transformants μg(-1)). Plasmids were rapidly lost under non-selective conditions with the frequency dependent on chromosomal element, thus recycling the acetamide- phenotype. The amdS plasmid was used to transform a set of clinical isolates resistant to a variety of antifungal drugs. All strains were successfully transformed to the acetamide+ phenotype at high frequency, confirming that this plasmid construct could be used as a simple dominant marker on virtually any strain. Gap repair experiments demonstrated that just as in Saccharomyces cerevisiae, gap repair functions efficiently inC. glabrata, suggesting that C. glabrata has numerous similarities toS. cerevisiae with regard to ease of molecular manipulation. The amdS system is inexpensive and efficient, and combined with existing C. glabrata plasmid elements, confers a high transformation frequency for C. glabrata with a phenotype that can be easily recycled.
authors.affiliation: The Department of Microbiology and Immunology, The University of Texas Health Science Center at San Antonio, San Antonio, TX 78229-3900, USA.
authors.forename: Jianmin
authors.initials: J
authors.lastname: Fu
authors.affiliation: The Department of Microbiology and Immunology, The University of Texas Health Science Center at San Antonio, San Antonio, TX 78229-3900, USA.
authors.forename: Morganne
authors.initials: M
authors.lastname: Blaylock
authors.affiliation: The Department of Microbiology and Immunology, The University of Texas Health Science Center at San Antonio, San Antonio, TX 78229-3900, USA.
authors.forename: Cameron F
authors.initials: CF
authors.lastname: Wickes
authors.affiliation: The Department of Microbiology and Immunology, The University of Texas Health Science Center at San Antonio, San Antonio, TX 78229-3900, USA.
authors.forename: William
authors.initials: W
authors.lastname: Welte
authors.affiliation: The Department of Microbiology and Immunology, The University of Texas Health Science Center at San Antonio, San Antonio, TX 78229-3900, USA.
authors.forename: Adrian
authors.initials: A
authors.lastname: Mehrtash
authors.affiliation: The Department of Pathology, The University of Texas Health Science Center at San Antonio, San Antonio, TX 78229-3900, USA.
authors.forename: Nathan
authors.initials: N
authors.lastname: Wiederhold
authors.affiliation: The Department of Microbiology and Immunology, The University of Texas Health Science Center at San Antonio, San Antonio, TX 78229-3900, USA wickes@uthscsa.edu.
authors.forename: Brian L
authors.initials: BL
authors.lastname: Wickes
mesh_terms: D000581:Amidohydrolases / Q000235:genetics* / Q000378:metabolism*; D001233:Aspergillus nidulans / Q000201:enzymology* / Q000235:genetics; D002176:Candida albicans / Q000235:genetics; D041221:Candida glabrata / Q000235:genetics* / Q000378:metabolism*; D003001:Cloning, Molecular; D005827:Genetics, Microbial / Q000379:methods*; D008967:Molecular Biology / Q000379:methods*; D010957:Plasmids; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid; D012641:Selection, Genetic; D014158:Transcription, Genetic; D014170:Transformation, Genetic
chemical_list: D011994:Recombinant Proteins; D000581:Amidohydrolases; C021445:acetamidase
keywords: ARS; CEN; acetamide; antifungal