title: Highly Efficient Electroporation-mediated Transformation into Edible Mushroom Flammulina velutipes.
abstract: In this study, we developed an efficient electroporation-mediated transformation system featuring Flammulina velutipes. The flammutoxin (ftx) gene of F. velutipes was isolated by reverse transcription-PCR. pFTXHg plasmid was constructed using the partial ftx gene (410 bp) along with the hygromycin B phosphotransferase gene (hygB) downstream of the glyceraldehydes-3-phosphate dehydrogenase (gpd) promoter. The plasmid was transformed into protoplasts of monokaryotic strain 4019-20 of F. velutipes by electroporation. High transformation efficiency was obtained with an electric-pulse of 1.25 kV/cm by using 177 transformants/µg of DNA in 1 × 10(7) protoplasts. PCR and Southern blot hybridization indicated that a single copy of the plasmid DNA was inserted at different locations in the F. velutipes genome by non-homologous recombination. Therefore, this transformation system could be used as a useful tool for gene function analysis of F. velutipes.
authors: Graduate School of Biotechnology & Information Technology, Hankyong National University, Ansung 456-749, Korea. ; Institute of Genetic Engineering, Hankyong National University, Ansung 456-749, Korea.
authors: Jong Kun
authors: JK
authors: Kim
authors: Young Jin
authors: YJ
authors: Park
authors: Won Sik
authors: WS
authors: Kong
authors: Hee Wan
authors: HW
authors: Kang
keywords: Electroporation; Flammulina velutipes; Flammutoxin gene; Hygromycin B gene; Protoplast
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