Details

Autor(en) / Beteiligte

• Sugui, Janyce A
• Chang, Yun C
• Kwon-Chung, K. J

Titel

Agrobacterium tumefaciens-Mediated Transformation of Aspergillus fumigatus: an Efficient Tool for Insertional Mutagenesis and Targeted Gene Disruption

Ist Teil von

• Applied and Environmental Microbiology, 2005-04, Vol.71 (4), p.1798-1802

Ort / Verlag

Washington, DC: American Society for Microbiology

Erscheinungsjahr

2005

Quelle

MEDLINE

Beschreibungen/Notizen

• Agrobacterium tumefaciens was used to transform Aspergillus fumigatus by either random or site-directed integration of transforming DNA (T-DNA). Random mutagenesis via Agrobacterium tumefaciens-mediated transformation (ATMT) was accomplished with T-DNA containing a hygromycin resistance cassette. Cocultivation of A. fumigatus conidia and Agrobacterium (1:10 ratio) for 48 h at 24°C resulted in high frequencies of transformation (>100 transformants/10⁷ conidia). The majority of transformants harbored a randomly integrated single copy of T-DNA and were mitotically stable. We chose alb1, a polyketide synthase gene, as the target gene for homologous integration because of the clear phenotype difference between the white colonies of [Delta]alb1 mutant strains and the bluish-green colonies of wild-type strains. ATMT with a T-DNA-containing alb1 disruption construct resulted in 66% albino transformants. Southern analysis revealed that 19 of the 20 randomly chosen albino transformants (95%) were disrupted by homologous recombination. These results suggest that ATMT is an efficient tool for transformation, random insertional mutagenesis, and gene disruption in A. fumigatus.