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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., 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. doi:10.1017/S0953756201005251 ISSN 0953-7562.
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Official URL: http://dx.doi.org/10.1017/S0953756201005251
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 | ||||
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Subjects: | S Agriculture > SB Plant culture | ||||
Divisions: | Faculty of Science, Engineering and Medicine > 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 | ||||
Dates: |
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Volume: | Vol.10 | ||||
Number: | No.1 | ||||
Page Range: | pp. 4-11 | ||||
DOI: | 10.1017/S0953756201005251 | ||||
Status: | Peer Reviewed | ||||
Access rights to Published version: | Open Access (Creative Commons) | ||||
Funder: | Royal Society (Great Britain), Great Britain. Ministry of Agriculture, Fisheries and Food (MAFF) |
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