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Current Genetics

, Volume 33, Issue 5, pp 378–385 | Cite as

Development of a homologous transformation system for the human pathogenic fungus Aspergillus fumigatus based on the pyrG gene encoding orotidine 5′′-monophosphate decarboxylase

  • Gerhard Weidner
  • Christophe d'Enfert
  • Andreas Koch
  • Pieternella C. Mol
  • A. A. Brakhage
ORIGINAL PAPER

Abstract

A homologous transformation system for the opportunistic fungal pathogen Aspergillus fumigatus was developed. It is based on the A. fumigatus pyrG gene, encoding orotidine 5′-monophosphate decarboxylase, which was cloned and sequenced. Transformation of both Aspergillus (Emericella) nidulans and A. fumigatus pyrG mutant strains by the use of protoplasts or electroporation established the functionality of the cloned gene. DNA sequencing of the A. fumigatus pyrG1 mutant allele revealed that it encodes a truncated, non-functional, PyrG protein. Transformation of an A. fumigatus pyrG1 mutant with a plasmid carrying the novel pyrG2 allele constructed by in vitro mutagenesis yielded prototrophic transformants following recombination between both mutation sites. Analysis of transformants carrying the entire plasmid showed that up to 45% of integration had occurred at the pyrG locus. This provides a tool to target defined genetic constructs at a specific locus in the A. fumigatus genome in order to study gene regulation and function.

Key wordsAspergillus fumigatus DNA-mediated transformation Orotidine 5′-monophosphate decarboxylase Electroporation 

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Copyright information

© Springer-Verlag Berlin Heidelberg 1998

Authors and Affiliations

  • Gerhard Weidner
    • 1
  • Christophe d'Enfert
    • 2
  • Andreas Koch
    • 1
  • Pieternella C. Mol
    • 2
  • A. A. Brakhage
    • 1
  1. 1.Lehrstuhl für Mikrobiologie, Universität München, Maria-Ward-Strasse 1 a, D-80638 München, Germany Fax: +49-89-17919862 e-mail: A.Brakhage@lrz.uni-muenchen.deDE
  2. 2.Laboratoire des Aspergillus, Institut Pasteur, 25, rue du Docteur Roux, F-75724 Paris Cedex 15, FranceFR

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