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Isolation, culture and division of olive (Olea europaea L.) protoplasts

Abstract

Protoplasts from Olea europaea L. have been compared in terms of their yield, viability, cell division and callus differentiation. Viable protoplasts were isolated from in vitro cultured leaves and cotyledons by an overnight incubation in an enzyme solution containing 1–1.5% driselase and 0.5M sucrose. This method allowed high yield of purified protoplasts, which floated and formed a dark green band at the meniscus, after centrifugation. Purified protoplasts were diluted to 3×104 protoplasts·ml−1 in culture medium. After cell wall regeneration, protoplasts gradually increased their volumes under appropriate conditions. The first divisions occurred during the second week in culture. Division efficiency ranged from 5.2 to 9.8% after 20 days in culture. Two weeks later visible microcolonies developed only from cotyledon protoplasts. After 6 weeks in culture, the microcalli were transferred to a solidified culture medium with 0.6% agarose, which induced active callus growth.

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Abbreviations

OM:

olive proliferation medium, Rugini 1984

Omg:

OM for the germination of olive embryos

OMr=OM:

for root induction

OMp=OM:

for protoplasts

OMc=OM:

for callus

BN:

Bourgin and Nitsch medium 1967

IBA:

indol-3-butyric acid

NAA:

naphthalene acetic acid

2,4-D:

dichlorophenoxyacetic acid.

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Author information

Correspondence to L. A. Cañas.

Additional information

Communicated by A. M. Boudet

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Cañas, L.A., Wyssmann, A.M. & Benbadis, M.C. Isolation, culture and division of olive (Olea europaea L.) protoplasts. Plant Cell Reports 6, 369–371 (1987). https://doi.org/10.1007/BF00269563

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Keywords

  • Sucrose
  • Cell Wall
  • Agarose
  • Cell Division
  • Solidify Culture Medium