Development of a simple and efficient transformation system for the basidiomycetous medicinal fungus Ganoderma lucidum |
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Authors: | Liang Shi Xing Fang Mengjiao Li Dashuai Mu Ang Ren Qi Tan Mingwen Zhao |
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Institution: | (1) Key Laboratory for Microbiological Engineering of the Agricultural Environment, Ministry of Agriculture, College of Life Sciences, Nanjing Agricultural University, No. 1 Weigang, Nanjing, 210095, Jiangsu, People’s Republic of China;(2) National Engineering Research Center of Edible Fungi, Shanghai Key Laboratory of Agricultural Genetics and Breeding, Institute of Edible Fungi, Shanghai Academy of Agricultural Science, Shanghai, 201106, China; |
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Abstract: | In this study, we report the development of a simple and efficient system for genetic transformation of the medicinal fungus
Ganoderma lucidum. Various parameters were optimized to obtain successful Agrobacterium tumefaciens-mediated transformation. Co-cultivation of bacteria and protoplast at a ratio of 1,000:1 at 25°C in medium containing 0.2 mM
acetosyringone was found to be the optimum condition for high efficiency transformation. Four plasmids, each carrying a different
promoter driving the expression of an antibiotic resistance marker, were tested. The construct carrying the Ganoderma lucidum glyceraldehyde-3-phosphate dehydrogenase (GPD) promoter showed good transformation efficiency, whereas constructs with the
GPD promoter from ascomycetes were ineffective. Our analysis showed that over 70% of the transformants tested remained mitotically
stable even after five successive rounds of subculturing. We were able to detect the expression of EGFP and GUS reporter genes
in the Ganoderma lucidum transformants by fluorescence imaging and histochemical staining assays respectively. Our results demonstrate a new transgenic
approach that will facilitate Ganoderma lucidum research. |
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