Agrobacterium tumefaciens-mediated Transformation of the Plant Pathogenic Fungus Rosellinia necatrix

作者：Kano Sanae; Kurita Takuma; Kanematsu Satoko; Morinaga Tsutomu^*

来源：Microbiology, 2011, 80(1): 82-88.

DOI：10.1134/S0026261711010103

• Agrobacterium tumefaciens
• hygromycin B
• Rosellinia necatrix
• T-DNA
• transformation

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 degrees 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.

• 出版日期2011-2