Plant regeneration from protoplasts isolated from embryogenic suspension cultured cells of Cinnamomum camphora L.
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An efficient and reproducible protocol is described for the regeneration of Cinnamomum camphora protoplasts isolated from cultured embryogenic suspension cells. Maximum protoplast yield (13.1±2.1×106/g FW) and viability (91.8±3.8%) were achieved using a mixture of 3% (w/v) cellulase Onozuka R10 and 3% (w/v) macerozyme Onozuka R10 in 12.7% (w/v) mannitol solution containing 0.12% (w/v) MES, 0.36% (w/v) CaCl2·2H2O, and 0.011% (w/v) NaH2PO4·2H2O. First divisions occurred 7–10 days following culture initiation. The highest division frequency (24.6±2.9%) and plating efficiency (6.88±0.8%) were obtained in liquid medium (MS) supplemented with 30 g l−1 sucrose, 0.7M glucose, 0.1 mg l−1 NAA, 1.0 mg l−1 BA, and 1.0 mg l−1 GA3. After somatic embryo induction and then shoot induction, the protoplast-derived embryos produced plantlets at an efficiency of 17.5%. Somatic embryos developed into well-rooted plants on MS medium supplemented with 1.0 mg l−1 3-indole butyric acid (IBA). Regenerated plants that transferred to soil have normal morphology.
KeywordsCinnamomum camphora L. Somatic embryogenesis Plant regeneration Protoplast culture
Cell protoplast wash;
3-Indole butyric acid;
2-(N-morpholino) Ethanesulphonic acid;
Murashige and Skoog medium;
Plant growth regulator;
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