Zhou Li-Hong, Wang Zheng-Xiang, Zhuge Jian
Research Center of Industrial Microbiology, Southern Yangtze University, Wuxi 214036, China.
Yi Chuan. 2006 Apr;28(4):479-85.
In order to facilitate the producer of polyketide pathway, four different transformation methods were tested and compared in an attempt to develop the genetic transformation system of Monascus sp. Using vector pBC-Hygro, the fungus was transformed to be hygromycin B-resistant, by conventional transformation as well as electroporation based on protoplast, electroporation based on germinated conidia, and restriction enzyme-mediated integration (REMI). Electroporation based on germinated conidia was found to be inappropriate for transforming Monascus sp. due to a low transformation frequency. The conventional transformation and electroporation technique based on protoplasts were thought not to be fit for transforming Monascus sp., due to a low stability of transformants though they yielded up to 135 transformants and 125 transformants per microgrammol/Lol/Le DNA, respectively. Transformant number was increased by 20-fold by REMI (2,500 transformants per microgrammol/Lol/Le DNA) and 70%-75% of them were stable. REMI technique would be very beneficial to the establishment of the genetic transformation system of Monascus sp.
为了促进聚酮化合物途径的生产者,测试并比较了四种不同的转化方法,试图开发红曲霉菌的遗传转化系统。使用载体pBC-Hygro,通过常规转化以及基于原生质体的电穿孔、基于萌发分生孢子的电穿孔和限制酶介导整合(REMI),将该真菌转化为对潮霉素B具有抗性。发现基于萌发分生孢子的电穿孔由于转化频率低而不适用于转化红曲霉菌。基于原生质体的常规转化和电穿孔技术虽然分别每微摩尔/升/升DNA产生多达135个转化体和125个转化体,但由于转化体稳定性低,被认为不适用于转化红曲霉菌。通过REMI,转化体数量增加了20倍(每微摩尔/升/升DNA有2500个转化体),其中70%-75%是稳定的。REMI技术将对红曲霉菌遗传转化系统的建立非常有益。