Liquid Culture Systems for in vitro Plant Propagation

Chapter 42
Tissue culture of Corydalis yanhusuo (Fumariaceae) and its
medicinal compound production from somatic embryoderived
plants
S. M. Nalawade 1 , A. P. Sagare 2 , C. L. Kuo 3 , S. F. Lo 4 , C. Y. Lee 1 , Y. L. Lee 5
& H. S. Tsay 1*
1
Institute of Biotechnology; Chaoyang University of Technology; 168, Gifeng E. Road,
Wufeng; Taichung 413; Taiwan.
*Corresp. author: tel: 886-4-2330-4920; fax: 2330-4921; e-mail: hstsay@mail.cyut.edu.tw
2
Division of Neurovascular Biology, Center for Aging and Developmental Biology, University
of Rochester Medical Center, Rochester, New York , USA.
3
Institute of Chinese Pharmaceutical Science, China Medical College, Taichung, Taiwan.
4
Department of Agronomy, Chiayi Agricultural Experiment Station, Agricultural Research
Institute, Chiayi, Taiwan.
5
Department of Food and Nutrition, Fu-Jen University, Taipei, Taiwan.
Abstract: An efficient method for regeneration of entire plants via somatic embryogenesis
in Corydalis yanhusuo using tuber-derived callus has been developed. Primary callus was
obtained by culturing tuber explants on Murashige and Skoog’s (MS) (1962) medium
supplemented with 2.0 mg l -1 N 6 -benzyladenine (BA) and 0.5 mg l -1 �-naphthaleneacetic acid
(NAA) in darkness for one month. Somatic embryos were induced by subculturing the
primary callus on MS medium supplemented with 0.5-4.0 mg l -1 BA, kinetin, or zeatin, within
2 weeks of culture in light. Rooting in the embryos with well developed cotyledonary leaves
was achieved by transferring them to half-strength liquid MS medium supplemented with
1.0 mg l -1 zeatin riboside for three weeks. Converted somatic embryos were cultured on halfstrength
MS medium supplemented with 6% (w/v) sucrose, and with 0.5-10.0 mg l -1 abscisic
acid (ABA), paclobutrazol, or ancymidol, 0.5-5.0 mg l -1 gibberellic acid (GA 3) and
15-100 mg l -1 polyethylene glycol (PEG) 4000 for further development of plantlets and tuber
formation in vitro. Recurrent somatic embryogenesis has been observed in the region at the
junction of cotyledon and root when the converted somatic embryos were cultured on MS
basal medium supplemented with 0-10 mg l -1 ABA. The use of liquid MS basal medium with
0.1 mg l -1 GA 3 enhanced embryo conversion within fifteen days of culture. Phenotypically
normal plants were recovered from converted somatic embryos and the plants showed welldeveloped
tuber formation and in vitro flowering after 4 months of culture. Also, the effects
of 0.5-5 mg l -1 ABA, paclobutrazol and 0.5-2 mg l -1 ancymidol, 0.5-5 mg l -1 GA 3 and
15-100 mg l -1 PEG 4000 supplemented in half-strength MS medium on the production of the
two major protoberberine-type alkaloids (D,L-tetrahydropalmatine and D-corydaline) by the
tubers of somatic embryo-derived plants of C. yanhusuo were examined. The high
557
A.K. Hvoslef-Eide and W. Preil (eds.), Liquid Culture Systems for in vitro Plant Propagation, 557–565.
© 2005 Springer. Printed in the Netherlands.