Transformation of Ipecac (Cephaelis ipecacuanha) with Agrobacterium rhizogenes

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 T_L-DNA was integrated into the plant genome without incorporation of the T_R-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)

Key words

Ipecac - Cephaelis ipecacuanha - Agrobacterium rhizogenes - transformation - isoquinoline alkaloids

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