Establishment of transformation system of Claviceps purpurea Cp-1 strain
10.16438/j.0513-4870.2016-0206
- VernacularTitle:麦角菌Cp-1菌株遗传转化体系的建立
- Author:
Wei-bo WANG
1
;
Jing-jing CHEN
1
;
Ping ZHU
1
Author Information
1. State Key Laboratory of Bioactive Substance and Function of Natural Medicines and Key Laboratory of Biosynthesis of Natural Products, National Health and Family Planning Commission of PRC, Institute of Materia Medica, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, China
- Publication Type:ORIGINAL ARTICLES
- Keywords:
Claviceps;
ergot alkaloid;
protoplast preparation;
genetic transformation
- From:
Acta Pharmaceutica Sinica
2016;51(5):828-
- CountryChina
- Language:Chinese
-
Abstract:
Claviceps pururea Cp-1 strain established in our lab is capable of producing variety of bioactive ergot alkaloids, and is broadly used by the pharmaceutical companies. To engineer the strain genetically for the production of specific ergot alkaloid, an effective transformation system must be set up first. However, the reported transformation system is not suitable for this strain due to different genetic backgrounds and the heterogeneity of Claviceps. Thus, in this paper, the hyphae of Cp-1 strain were used to prepare protoplasts by lywallzyme. The formation of protoplasts was investigated under different concentrations and incubation time of enzyme. The strain was tested for sensitivity to several antibiotics at different concentrations. Finally, the genetic transformation system of Cp-1 strain was established. The results suggest that protoplasts were formed efficiently by using 1% lywallzyme at 25℃ for 2 h. Transformants were obtained by PEG mediated protoplast transformation of Cp-1 strain with plasmid pAN7-1, using 1.5 mg·mL-1 hygromycin B as the selective marker. The exogenous gene in the plasmid pAN7-1 was integrated into the genome of Cp-1 strain transformant as demonstrated by PCR result. This study laid an important foundation for genetic manipulation of Cp-1 strain.
