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A reverse genetic approach for generating gene replacement mutants in Ustilago maydis

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An Erratum to this article was published on 29 September 2004

Abstract

We describe a versatile strategy for generating gene replacement mutants in the phytopathogenic fungus Ustilago maydis. The system includes the choice of 32 different insertion cassettes for genetic engineering purposes, such as gene disruption and more sophisticated insertions of reporter genes, heterologous promoters or combinations of the two. PCR-amplified flanking sequences needed for homologous recombination are ligated to the respective insertion cassettes via Sfi I sites. As proof of principle we generated two replacement mutants in which the endogenous promoter of the pheromone gene mfa1 drives expression of the Green Fluorescent Protein gene (gfp). Simultaneously, expression of the mfa1 ORF is controlled either by the carbon source-regulated crg1 promoter or the nitrogen source-regulated nar1 promoter. In both cases gfp expression was pheromone-inducible and pheromone expression was only detected when the heterologous promoters were active.

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Fig. 1A, B
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Acknowledgements

We acknowledge Dr. R. Kahmann for critically reading the manuscript, and thank J. Hohenner, S. Hester and P. Roth for excellent technical assistance. This work was supported by the DFG through Sonderforschungsbereich 369 and by funding from Bayer CropScience

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Author notes

  1. A. Brachmann

    Present address: National Institute for Diabetes and Digestive and Kidney Diseases, NIH, 8 Center Drive, Bethesda, MD 20892-0830, USA

Authors and Affiliations

  1. Department of Organismic Interactions, Max Planck Institute for Terrestrial Microbiology, Karl-von-Frisch-Strasse, 35043, Marburg, Germany

    A. Brachmann, J. König, C. Julius & M. Feldbrügge

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  1. A. Brachmann
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  2. J. König
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Correspondence to M. Feldbrügge.

Additional information

Communicated by G. Jürgens

The first two authors contributed equally to this work

An erratum to this article is available at http://dx.doi.org/10.1007/s00438-004-1067-8.

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Brachmann, A., König, J., Julius, C. et al. A reverse genetic approach for generating gene replacement mutants in Ustilago maydis. Mol Genet Genomics 272, 216–226 (2004). https://doi.org/10.1007/s00438-004-1047-z

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  • DOI: https://doi.org/10.1007/s00438-004-1047-z

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