Abstract
A plant regeneration system from the isolated protoplasts of Echinacea purpurea L. using an alginate solid/liquid culture is described in the chapter. Viable protoplasts were isolated from 100 mg of young leaves of 4-wk-old seedlings in an isolation mixture containing 1.0% cellulase Onozuka R-10, 0.5% pectinase, and 0.3 mol/L mannitol. After isolation and purification, the mesophyll protoplasts were embedded into 0.6% Na-alginate at the density 1 × 10−5 mL and cultured in modified Murashige and Skoog (MS) culture medium supplemented with 0.3 mol/L sucrose, 2.5 µmol/L benzylaminopurine (BA), and 5.0 µmol/L 2,4-dichlorophenoxyacetic acid (2,4-D). The visible colonies were present after 4 wk of culture. The protoplast-derived clones were transferred onto gellan gum-solidified basal medium supplemented with 1.0 µmol/L BA and 2.0 µmol/L indole-3-butyric acid (IBA) and formed compact and green calli. Shoot development was achieved by subculturing the calli onto the same basal medium supplemented with 5.0 µmol/L BA and 2.0 µmol/L IBA. Further subculture onto basal medium resulted in the regeneration of complete plantlets.
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Acknowledgments
The financial support of the Natural Sciences and Engineering Research Council of Canada and the Ontario Ministry of Food Agriculture is gratefully acknowledged.
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Pan, Zg., Liu, Cz., Murch, S.J., Saxena, P.K. (2006). Isolation, Culture, and Plant Regeneration From Echinacea purpurea Protoplasts. In: Loyola-Vargas, V.M., Vázquez-Flota, F. (eds) Plant Cell Culture Protocols. Methods in Molecular Biology™, vol 318. Humana Press. https://doi.org/10.1385/1-59259-959-1:211
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DOI: https://doi.org/10.1385/1-59259-959-1:211
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