High-frequency Direct Somatic Embryogenesis and Plantlet Regeneration from Leaves Derived from In Vitro-Germinated Seedlings of a Coffea arabica Hybrid Cultivar

in HortScience

Breeding work carried out during the period 1971–85 by the Coffee Research Institute, Ruiru, Kenya resulted in the release of a new improved hybrid Coffea arabica named Ruiru 11. The cultivar combines resistance to coffee berry disease (CBD) and leaf rust, with high yield and good cup quality attributes. The propagation by F1 hybrid seeds production, cuttings, and tip grafting do not produce enough planting materials. There was a need to explore alternative methods and tissue culture offers potential options. The objective of the study was to evaluate the effect of explant sources and cytokinins on induction and regeneration of somatic embryos. Eight different explants were cultured on half-strength Murashige and Skoog (MS) medium supplemented with 10 µm benzylaminopurine (BAP). The effect of kinetin, N6-(2-isopentyl) adenine (2iP) evaluated at (0, 0.5, 5, or 25 µm) or thidiazuron (TDZ) (0, 0.5, 1.0, or 5 µm) added in separate experiments was also evaluated. The percentage of embryogenic cultures and the numbers of embryos per explant were determined after 3 months’ culture. The explant type had a significant effect (P > 0.05) on the induction of somatic embryos. Explants from in vitro-germinated seedlings produced the highest embryogenic cultures (90%) and the highest mean number of embryos (19.36) per explant. Cytokinins strongly enhanced induction and regeneration of somatic embryos. TDZ at 1 µm produced the highest embryogenic cultures (100%) and the highest mean number of embryos (24.2). The embryos were germinated on half-strength MS medium without any hormones. A high (98%) survival rate of the regenerated plantlets was recorded over all the treatments in the greenhouse. This is the first report on induction of high-frequency direct somatic embryos from coffee juvenile tissues. This is of great significance in tissue culture and indeed molecular biology manipulations because it allows regeneration of coffee from several explants.

Contributor Notes

We wish to acknowledge and thank Prof Esther Kahangi of the Jomo Kenyatta University of Agriculture and Technology for her contributions to this article.

Corresponding author. E-mail: j.kahia@cgiar.org.

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Article Figures

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    Effect of different explants types on induction and regeneration of somatic embryos. (A) The frequency of somatic embryo regeneration. (B) The mean number of embryos. 1, Leaves from in vitro-germinated seedlings; 2, leaves from 6-month-old seedlings; 3, hypocotyls of embryo 4, leaves from in vitro-propagated somatic embryos; 5, lateral roots of somatic embryos; 6, primary roots of embryos; 7, cotyledonary leaves of embryos; and 8, leaves from field-grown trees.

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    (A) Different stages of coffee somatic embryos derived from the media supplemented with thidiazuron 1 µm. (a) Globular, (b) heart-shaped, (c) torpedo, and (d) early cotyledonary stage in a single culture after 90 d in culture. (B) Plantlet with well-developed shoot and root ready for weaning.

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    In vitro-propagated coffee plants in the greenhouse. (A) Weaning of coffee plantlets. (B) Coffee plantlet ready for planting in the field.

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