A highly efficient polyethylene glycol-mediated transformation method for mushrooms.
A highly efficient transformation system mediated by polyethylene glycol was developed for the cultivated mushroom Pleurotus ostreatus. Eighty to 180 integrative and stable-resistant colonies appeared per mug of DNA per 10(7) viable protoplasts in a transformation experiment with the hygromycin B phosphotransferase gene (hph), which is about 40-1800 times higher than that previously reported in P. ostreatus. One hundred to 150 transformants emitting green fluorescence were observed per mug of DNA per 10(7) viable protoplasts in a transformation with the green fluorescent protein gene, but green fluorescence disappeared 30 h after transformation, suggesting that the green fluorescent protein gene was only transiently expressed in P. ostreatus. Plasmid pAN7-1 was also transferred into two important cultivated mushrooms, Ganoderma lucidum and Lentinus edodes, and 120-150 and 85-100 transformants per mug of DNA per 10(7) viable protoplasts were obtained, respectively, which is seven to 38 times and 24-28 times greater than previously reported. These data indicate that this new polyethylene glycol-mediated transformation procedure is highly efficient for mushrooms, and could be a useful tool in mushroom improvement by gene engineering.
Li
Gang
G
The Key Laboratory of Gene Engineering of Ministry of Education, Sun Yat-sen University, Guangzhou, China.
Li
Ruixue
R
Liu
Qiuyun
Q
Wang
Qiang
Q
Chen
Min
M
Li
Baojian
B
D025141:Drug Resistance, Fungal / Q000235:genetics; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000032:analysis / Q000235:genetics; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000235:genetics; D010957:Plasmids / Q000235:genetics; D020076:Pleurotus / Q000235:genetics*; D011092:Polyethylene Glycols / Q000494:pharmacology*; D011523:Protoplasts; D020940:Reishi / Q000235:genetics*; D020942:Shiitake Mushrooms / Q000235:genetics*; D014170:Transformation, Genetic
D049452:Green Fluorescent Proteins; D011092:Polyethylene Glycols; D017853:Phosphotransferases (Alcohol Group Acceptor); C063242:hygromycin-B kinase