Highly Efficient Electroporation-mediated Transformation into Edible Mushroom Flammulina velutipes
Mycobiology
;
: 331-335, 2010.
Article
in English
| WPRIM
| ID: wpr-729904
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/microg of DNA in 1 x 107 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.
Full text:
Available
Index:
WPRIM (Western Pacific)
Main subject:
Oxidoreductases
/
Plasmids
/
Protoplasts
/
Recombination, Genetic
/
Sprains and Strains
/
DNA
/
Fungal Proteins
/
Hygromycin B
/
Chimera
/
Blotting, Southern
Language:
English
Journal:
Mycobiology
Year:
2010
Type:
Article
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