Chinese Bulletin of Botany ›› 2019, Vol. 54 ›› Issue (1): 72-80.DOI: 10.11983/CBB18118

• TECHNIQUE AND METHOD • Previous Articles     Next Articles

Establishment of a Genetic Transformation System for Dioscorea opposita Using Microtuber

Junhua Li1,2,3,,Shiyu Liu1,,Chenglong Li1,Linlin Han1,Yahui Dong1,Xiaoli Zhang1,2,3,Xiting Zhao1,2,3,Mingjun Li1,2,3,*()   

  1. 1 College of Life Science, Henan Normal University, Xinxiang 453007, China
    2 Engineering Technology Research Center of Nursing and Utilization of Genuine Chinese Crude Drugs of Henan Province, Xinxiang 453007, China
    3 Engineering Laboratory of Green Medicinal Material Biotechnology of Henan Province, Xinxiang 453007, China
  • Received:2018-05-15 Accepted:2018-09-13 Online:2019-01-30 Published:2019-07-31
  • Contact: Junhua Li,Shiyu Liu,Mingjun Li

Abstract: Genetic transformation system is the foundation for gene function study and genetic improvement of Dioscorea opposita. In this study, we established an Agrobacterium-mediated genetic transformation system using microtuber in D. opposita cv. ‘Tiegun’. The transformation was performed with the DoSERK2 gene overexpression plasmid pCAMBIA1301-DoSERK2 that harboring the GUS reporter gene and the hygromycin (Hyg) resistence gene as the selectable marker, and the gene-silencing vector pART27-DoSERK2 that containing kanamycin (Kan) resistence marker. The optimal concentration of timentin (Tim) for Agrobacterium counter selection was 500 mg·L -1. For shoot regeneration and rooting, the optimal concentrations of Hyg were 15 mg·L -1 and 20 mg·L -1, respectively, and the optimal concentration of Kan were 120 mg·L -1 and 160 mg·L -1, respectively. Results of PCR analysis and GUS histochemical staining indicated that the exogenous genes had been integrated into the genome of transgenic plants, and are expressed in the plants. Our study provides a genetic transformation system in D. opposita cv. ‘Tiegun’, which has the advantage that explants are easy available. This method has reference value for the genetic transformation of other varieties of D. opposita.

Key words: Agrobacterium tumefaciens, genetic transformation, microtuber, Dioscorea opposita cv. ‘Tiegun’