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Efficient micropropagation of Ensete ventricosum applying meristem wounding: a three-step protocol

  • Cell Biology and Morphogenesis
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Abstract

A highly efficient three-step protocol for in vitro propagation of Ensete ventricosum (enset) was developed that consisted of initiation, bud proliferation, and shoot elongation and rooting stages. At the initiation stage, it was crucial to use shoot tips (5–8 mm) with subtending corm tissues as explants to obtain growth. The addition of 0.5–1% (w/v) activated charcoal to the medium was essential to prevent phenol exudation which otherwise leads to the loss of cultures. During the bud proliferation stage, modified MS macronutrients and micronutrients together with a combination of cytokinins (1.6 μM naphthaleneacetic acid, 4.4 μM 6-benzylaminopurine, 23.2 μM kinetin, 22.6 μM N6 2-isopentyladenine) was used. This novel composition of macronutrients was based on the analysis of leaf nutrient content of glasshouse-grown enset sprouts. Multiple bud formation on the enlarged corm tissue was induced only when the meristem region was wounded before transfer to the bud proliferation medium. Up to 75 healthy shoots per explant were produced, whereas unwounded explants produced, only one to two shoots per explant. A third stage with a low concentration of cytokinin enabled shoot elongation as well as root development. The plantlets were acclimatized with 100% success and they showed no apparent phenotypical deviation.

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Fig. 1a–i

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Abbreviations

BAP:

6-Benzylaminopurine

IBA:

Indole-3-butyric acid

DW:

Dry weight

EV:

Ensete ventricosum medium

2-iP N6:

2-Isopentyl adenine

NAA:

α-Naphthaleneacetic acid

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Acknowledgements

The financial support obtained from Swedish International Development Authority (SIDA), co-ordinated under the Stockholm Environment Institute (SEI) is gratefully acknowledged. Our deep gratitude goes to assistant professor Bengt Bengtsson and Mr. Sven Svennsson for their help during the analysis of mineral nutrients and to Mrs. Annelie Ahlman and Dr. Li-Hua Zhu for their technical advice during the laboratory work. Dr. Salla Martilla and Mrs. Kerstin Brismar are acknowledged for their help with light and scanning microscopy and Prof. Waheeb Heneen for his advice during this work. We are also grateful to the staff of the Biotron and the glasshouse for looking after our plants. Finally. we would like to express our thanks to Agida Incorporated for performing the pathogen testing of our samples.

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Authors and Affiliations

  1. Department of Crop Science, Swedish University of Agricultural Sciences, P.O. Box 44, 23053, Alnarp, Sweden

    G. Birmeta & M. Welander

  2. Department of Biology, Addis Ababa University, P.O. Box 1176, Addis Ababa, Ethiopia

    G. Birmeta

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  1. G. Birmeta
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  2. M. Welander
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Correspondence to G. Birmeta.

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Communicated by E. Heberle-Bors

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Birmeta, G., Welander, M. Efficient micropropagation of Ensete ventricosum applying meristem wounding: a three-step protocol. Plant Cell Rep 23, 277–283 (2004). https://doi.org/10.1007/s00299-004-0832-9

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  • DOI: https://doi.org/10.1007/s00299-004-0832-9

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