Li Gang, Li Ruixue, Liu Qiuyun, Wang Qiang, Chen Min, Li Baojian
The Key Laboratory of Gene Engineering of Ministry of Education, Sun Yat-sen University, Guangzhou, China.
FEMS Microbiol Lett. 2006 Mar;256(2):203-8. doi: 10.1111/j.1574-6968.2006.00110.x.
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.
开发了一种由聚乙二醇介导的高效转化系统，用于栽培蘑菇平菇。在用潮霉素B磷酸转移酶基因（hph）进行的转化实验中，每10⁷个活原生质体每微克DNA出现80至180个整合且稳定抗性的菌落，这比平菇先前报道的效率高约40至1800倍。在用绿色荧光蛋白基因进行的转化中，每10⁷个活原生质体每微克DNA观察到100至150个发出绿色荧光的转化体，但转化30小时后绿色荧光消失，这表明绿色荧光蛋白基因仅在平菇中瞬时表达。质粒pAN7-1也被转入两种重要的栽培蘑菇，灵芝和香菇，每10⁷个活原生质体每微克DNA分别获得120至150个和85至100个转化体，这分别比先前报道的高7至38倍和24至28倍。这些数据表明，这种新的聚乙二醇介导的转化方法对蘑菇具有高效性，并且可能是通过基因工程改良蘑菇的有用工具。
