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PEG-mediated and Agrobacterium-mediated transformation in the mycopathogen Verticillium fungicola

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Amey, Richard C., Athey-Pollard, Anna, Burns, C. (Claire), Mills, P. R. (Peter R.), Bailey, Andy M. and Foster, Gary D. . (2002) PEG-mediated and Agrobacterium-mediated transformation in the mycopathogen Verticillium fungicola. Mycological Research, Vol.10 (No.1). pp. 4-11. ISSN 0953-7562

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Abstract

Verticillium fungicola, a severe mycopathogen of the cultivated mushroom Agaricus bisporus, was successfully transformed using both PEG-mediated and Agrobacterium-mediated techniques. PEG-mediated co-transformation was successful with hygromycin B resistance (hph), uidA (β-glucuronidase GUS), and green fluorescent protein (GFP) genes. Agrobacterium-mediated transformation was successful with the hph gene. Transformation frequencies of up to 102 transformants per μg DNA and 4068 transformants per 105 conidia were obtained for PEG-mediated and Agrobacterium-mediated transformation respectively. Expression of integrated genes in co-transformants was stable after 18 months of successive sub-culturing on non-selective medium, and following storage at -80 °C in glycerol. Molecular analysis of PEG-mediated transformants showed integration of the transforming genes into the target genome. Molecular analysis of Agrobacterium-mediated transformants showed integration of transforming DNA as single copies within the target genome. Co-transformants exhibited symptoms of disease in inoculation experiments and were at least as virulent as the wild-type fungus. GFP and GUS expression were observed in-vivo with the GFP-tagged strain showing great potential as a tool in epidemiological and host-pathogen interaction studies. The development of transformation systems for V. fungicola will allow in-depth molecular studies of the interaction of this organism with A. bisporus.

Item Type: Journal Article
Subjects: S Agriculture > SB Plant culture
Divisions: Faculty of Science > Life Sciences (2010- ) > Warwick HRI (2004-2010)
Library of Congress Subject Headings (LCSH): Verticillium, Fungi imperfecti, Cultivated mushroom -- Diseases and pests, Agrobacterium, Fungi -- Genetics
Journal or Publication Title: Mycological Research
Publisher: Cambridge University Press
ISSN: 0953-7562
Official Date: January 2002
Volume: Vol.10
Number: No.1
Page Range: pp. 4-11
Identification Number: 10.1017/S0953756201005251
Status: Peer Reviewed
Access rights to Published version: Open Access
Funder: Royal Society (Great Britain), Great Britain. Ministry of Agriculture, Fisheries and Food (MAFF)
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URI: http://wrap.warwick.ac.uk/id/eprint/789

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