Genetic transformation of the symbiotic basidiomycete fungus Hebeloma cylindrosporum |
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Authors: | roland Marmeisse Gilles Gay Jean-Claude Debaud Lorna A Casselton |
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Institution: | (1) School of Biological Sciences, Queen Mary and Westfield College, Mile End Road, E1 4NS London, UK;(2) Laboratoire CNRS d'Ecologie Microbienne du Sol, Université Lyon 1, Bât. 405, 43 Boulevard du 11 Novembre, F-69622 Villeurbanne Cedex, France;(3) Present address: Department of Genetics, Agricultural University, Dreijenlaan 2, NL-6703 HA Wageningen, The Netherlands;(4) Present address: Department of Plant Sciences, University of Oxford, South Parks Road, OX1 3RB Oxford, UK |
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Abstract: | Summary The pAN7.1 plasmid containing the E. coli hygromycin B phosphotransferase gene was used to transform protoplasts of the ectomycorrhizal fungus Hebeloma cylindrosporum. Hygromycin-resistant transformants were selected at a frequency of one to five per g of transforming DNA. Southern blot analyses revealed multiple copy integration of the transforming plasmid into the genome. The selection system was used to introduce other genes of interest by co-transformation. Two plasmids, one containing tryptophan biosynthesis genes and the other the NADP-glutamate dehydrogenase gene from the saprophytic basidiomycete Coprinus cinereus, were successfully introduced into the H. cylindrosporum genome with up to 70% efficiency of co-transformation. The hygromycin resistance phenotype was stably maintained during growth of transformants on hygromycinfree medium. All tranformants retained their ability to form mycorrhizae with the habitual host plant Pinus pinaster, making them suitable for future physiological studies. |
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Keywords: | Transformation Hygromycin Mycorrhizal fungi Hebeloma |
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