Planta Med 2003; 69(11): 1018-1023
DOI: 10.1055/s-2003-45149
Original Paper
Physiology, in vitro Biotechnology
© Georg Thieme Verlag Stuttgart · New York

Transformation of Ipecac (Cephaelis ipecacuanha) with Agrobacterium rhizogenes

Kayo Yoshimatsu1 , Koichiro Shimomura1, 2 , Mami Yamazaki3 , Kazuki Saito3 , Fumiyuki Kiuchi1
  • 1Tsukuba Medicinal Plant Research Station, National Institute of Health Sciences, Tsukuba, Ibaraki, Japan
  • 2Present address: Faculty of Life Sciences, Toyo University, Itakura-machi, 0ura-gun, Gunma, Japan
  • 3Department of Molecular Biology and Biotechnology, Graduate School of Pharmaceutical Sciences, Chiba University, Chiba, Japan
This work was supported in part by Special Cooperation Funds for Promoting Science and Technology (Basic Research Core System) from the Science and Technology Agency in Japan
Further Information

Publication History

Received: May 7, 2003

Accepted: August 16, 2003

Publication Date:
09 January 2004 (online)

Abstract

Transformed root cultures of ipecac (Cephaelis ipecacuanha A. Richard), one of the recalcitrant woody plant species for Agrobacterium-mediated transformation, were established by co-culturing of in vitro petiole segments with Agrobacterium rhizogenes ATCC 15 834. Southern blot analysis of the established roots revealed that only the TL-DNA was integrated into the plant genome without incorporation of the TR-DNA. The transformed roots grew slowly on phytohormone-free solid medium and adventitious shoots were regenerated after over 6 months of culture on HF, half-strength Murashige and Skoog (1/2 MS) medium in the dark. The individually separated transformed shoots developed into plantlets on phytohormone-free solid medium at 25 °C under 16 h/day light, and the plants demonstrated wider leaves, shorter internodes and vigorous root growth compared to non-transformed plants. Effects of basal media and auxins on the growth and the ipecac alkaloid production of the transformed roots were investigated either under light or in the dark. The roots cultured in the dark grew well in Gamborg B5 (B5) liquid medium containing 0.5 mg/L IBA and yielded 112 mg/L of cephaeline and 14 mg/L emetine after 8 weeks of culture.

Abbreviations

B5: Gamborg B5 (1968)

HF:phytohormone-free

IAA:indole-3-acetic acid

IBA:indole-3-butyric acid

MS:Murashige and Skoog (1962)

1/2 MS:a half strength Murashige and Skoog

NAA:α-naphthaleneacetic acid

WP:Woody Plant (1980)

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Kayo Yoshimatsu

Tsukuba Medicinal Plant Research Station

National Institute of Health Sciences

1 Hachimandai

Tsukuba

Ibaraki 305-0843

Japan

Phone: +81-29-838-0573

Fax: +81-29-838-0575

Email: yoshimat@nihs.go.jp