Isolation and culture of suspension-derived protoplasts ofBeta vulgaris L. |
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Authors: | A Majewska-Sawka H Nakashima K Mori |
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Institution: | 1. Institute for Plant Breeding and Acclimatization, Bydgoszcz, Poland 2. Faculty of Agriculture, Hokkaido University, Sapporo, Japan
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Abstract: | Sugar beet protoplasts (Beta vulgaris L.) were isolated from hypocotyl-derived suspension cells and cultured on modified Murashige and Skoog medium supplemented
with 5 μM naphthaleneacetic acid (NAA) and 2 μM 6-benzyl-aminopurine (BAP). Protoplasts were plated at a density 1.0–1.5×105 cm−3 and incubated in either liquid medium or in medium solidified by 1.2% agarose, at 25°C in the dark. Comparison of two methods
of culture unequivocally showed the second to be superior. Immobilizing the protoplast in agarose proved to be essential for
obtaining sustained protoplast division and reproducible colony formation. The plating efficiency after two weeks of culture,
expressed as the percentage of protoplasts which developed to form colonies, reached 40%. Subsequent subcultures of protoplast-derived
callus to regeneration media with different concentrations of BAP (5 μM, 10 μM, 20 μM, 30 μM) resulted in very good callus
proliferation at the three lowest concentrations, although organogenesis was not achieved. |
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