Plant Cell, Tissue and Organ Culture

, Volume 27, Issue 3, pp 231–241

High plating efficiency and plant regeneration frequency in low density protoplast cultures derived from an embryogenic Brassica napus cell suspension

  • Daina H. Simmonds
  • Nancy E. Long
  • Wilfred A. Keller


Protoplasts were isolated from an embryogenic cell suspension culture derived from microspores of Brassica napus cv. Jet Neuf. Protoplast yield varied with the cell suspension growth medium. Optimization of protoplast plating density, manipulation of culture medium, carbon source and medium matrix, and inclusion of Ficoll resulted in protoplast plating efficiencies close to 30%. Placement of the protoplasts close to the gas interface contributed greatly to the elevated plating efficiency. Low density cultures could be induced to regenerate calli at optimum plating efficiencies if grown in the presence of nurse culture. This is of great advantage for manipulation of individual protoplasts or for microinjection. Plants were regenerated directly from the cell suspension or from the protoplast cultures.

Key words

agarose Ficoll filter paper gas-exchange micromanipulation 





2,4-dichlorophenoxyacetic acid


indole-3-acetic acid


naphthaleneacetic acid


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Copyright information

© Kluwer Academic Publishers 1991

Authors and Affiliations

  • Daina H. Simmonds
    • 1
  • Nancy E. Long
    • 1
  • Wilfred A. Keller
    • 1
  1. 1.Plant Biotechnology Program, Plant Research CentreAgriculture CanadaOttawa(Canada)
  2. 2.Plant Biotechnology InstituteNational Research Council of CanadaSaskatoon

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