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Establishment of Transgenic Acceptor by Indirect Somatic Embryogenesis Regeneration and Transformation of CmTGA1 Gene in Chrysanthemum morifolium cv. ‘Huaihuang’

Xiting Zhao1, 2, 3*, Liwei Jiang1, Miao Wang1, Yuting Zhu1, Wenfang Zhang1, Mingjun Li1, 2, 3*   

  1. 1College of Life Sciences Henan Normal University, Xinxiang 453007, China; 

    2Engineering Technology Research Center of Nursing and Utilization of Genuine Chinese Crude Drugs of Henan Province University, Xinxiang 453007, China; 

    3Henan Province Engineering Laboratory of Green Medicinal Plant Biotechnology, Xinxiang 453007, China
  • Received:2015-10-09 Revised:2016-03-07 Online:2016-08-05 Published:2016-07-01
  • Contact: Xiting Zhao,Mingjun Li E-mail:zhaoxt0411@126.com; limingjun2002@263.net

Abstract:

The transgenic acceptor regeneration system and the genetic transformation system are important in the plant transgenic manipulation. This study explored the optimal medium of embryoid induction and the optimal antibiotic selection concentrations of shoot and root regeneration of Chrysanthemum morifolium cv. ‘Huaihuang’. CmTGA1, a disease- resistant gene, was transformed into ‘Huaihuang’ by using homeosis and regenerated new lines. Explants were cultured for 15 d on MS basal medium supplemented with 1.5 mg·L−1 IAA, 0.5 mg·L−1 6-BA and 1.0 mg·L−1 2,4-D, then transferred to the meristematic medium without 2,4-D and cultured for 8−10 weeks, finally the regeneration bud was induced for 2 to 3 weeks on the developed medium. 86% of explants gained regeneration buds from embryoids. The selected concentration of hygromycin B was 5.0 mg·L−1 for shoot regeneration and 4.5 mg·L−1 for rooting. The integration of CmTGA1 into the genome of the transformation lines was confirmed by semi-quantitative PCR and real-time quantitative PCR. Finally, the transgenic acceptor regeneration system with indirect somatic embryogenesis was established successfully for Huaihuang, and this technology may lay the foundation for solving some production problems such as serious disease infection and germplasm degradation by genetic engineering methods.

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