Krappman, Sven and Bayram, Ozgur and Braus, Gerhard H.
(2005)
Deletion and Allelic Exchange of the Aspergillus fumigatus veA Locus
via a Novel Recyclable Marker Module.
Eukaryotic Cell, 4 (7).
pp. 1298-1307.
ISSN 1535-9778
Abstract
Detailed evaluation of gene functions in an asexual fungus requires advanced methods of molecular biology.
For the generation of targeted gene deletions in the opportunistic pathogen Aspergillus fumigatus we designed
a novel blaster module allowing dominant selection of transformants due to resistance to phleomycin as well
as dominant (counter)selection of a Cre recombinase-mediated marker excision event. For validation purposes
we have deleted the A. fumigatus pabaA gene in a wild-type isolate by making use of this cassette. The resulting
pabaA::loxP strain served as the recipient for subsequent targeting of the velvet locus. Homologous reconstitution
of the deleted gene was performed by an allele whose expression is driven in a nitrogen source-dependent
manner, as validated by Northern analyses. Overexpression of the veA locus in A. fumigatus does not result in
any obvious phenotype, whereas the sporulation capacities of the veA null mutant are reduced on nitratecontaining
medium, a phenotype that is completely restored in the reconstituted strain.
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