1. Agrobacterium tumefaciens-mediated transformation of the plant pathogenic fungus Rosellinia necatrix.
- Author
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Kano, Sanae, Kurita, Takuma, Kanematsu, Satoko, and Morinaga, Tsutomu
- Subjects
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AGROBACTERIUM tumefaciens , *ASCOMYCETES , *PLANT species , *MYCOSES , *PHYTOPATHOGENIC microorganisms , *FILAMENTOUS fungi - Abstract
Rosellinia necatrix is a soil-borne root pathogen affecting a wide range of commercially important plant species. The mycelium of R. necatrix was transformed to hygromycin B resistance by an Agrobacterium tumefaciens-mediated transformation system using a binary plasmid vector containing the hygromycin B phosphotransferase ( hph) gene controlled by the heterologous fungal Aspergillus nidulans P- gpd (glyceraldehyde 3-phosphate dehydrogenase) promoter and the trpC terminator. Co-cultivation of R. necatrix strain W1015 and A. tumefaciens strain AGL-1 at 25°C using the binary vector pAN26-CB1300, which contained the hygromycin B resistance cassette based on pAN26 and pCAMBIA1300, resulted in high frequencies of transformation. The presence of the hph gene in the transformants was detected by PCR, and single-copy integration of the marker gene was demonstrated by Southern b lot analy s is. This report of an Agrobacterium-mediated transformation method should allow the development of T-DNA tagging as a system for insertional mutagenesis in R. necatrix and provide a simple and reliable method for genetic manipulation. [ABSTRACT FROM AUTHOR]
- Published
- 2011
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