Abstract
Agrobacterium tumefaciens was used to stably transform the entomopathogenic deuteromycete Beauveria bassiana to hygromycin B resistance by integration of the hph gene of Escherichia coli into the fungal genome. The transformation protocol was optimized to generate a library of insertion mutants of Beauveria. Transformation frequencies around 10−4 and suppression of background growth were achieved. Over 90% of the AIM mutants investigated contained single-copy T-DNA integrations at different chromosomal locations. Integrated T-DNAs were re-isolated from ten transformants by a marker rescue approach. When the sequences flanking these T-DNAs were compared with the corresponding locations of the wild-type genome, truncations of T-DNA borders were found to be common, while none of the sites of integration had suffered deletion or rearrangement. Thus, AIM can be considered a promising tool for insertional mutagenesis studies of entomopathogenic filamentous fungi.
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Acknowledgements
We are highly indebted to Paul Bundock for providing the A. tumefaciens strain used in this study and to Seogchan Kang for vector pBHt2 and some useful hints. The work presented in this publication was financially supported by the Deutsche Forschungsgemeinschaft (DFG) in the form of research grants and a scholarship from the DFG-Graduiertenkolleg for H.W. Both G.V.M. and A.L. received grants from the Deutsche Akademische Austauschdienst (DAAD).
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Communicated by J. Heitman
A. Leclerque and H. Wan contributed equally to this work
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Leclerque, A., Wan, H., Abschütz, A. et al. Agrobacterium-mediated insertional mutagenesis (AIM) of the entomopathogenic fungus Beauveria bassiana . Curr Genet 45, 111–119 (2004). https://doi.org/10.1007/s00294-003-0468-2
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DOI: https://doi.org/10.1007/s00294-003-0468-2