Normalizing sweet orange (C. sinensis(L.) osbeck) somatic embryogenesis with semi-permeable membranes

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SummaryDevelopment of citrus somatic embryos initiated from embryogenic callus generally results in abnormal morphogenesis of somatic embryos. To normalize development, glycerol-induced globular-stage somatic embryos of sweet orange [C. sinensis (L.) Osbeck cv. ‘Hamlin'] were cultured on 6000-8000 MW cutoff cellulose acetate, >400 000 MW cutoff cellulose acetate, nitrocellulose, polyvinylidene fluoride (PVDF), cellulose filter paper, or positive or neutral charged nylon membranes. Only the two cellulose acetate membranes resulted in the development of normal, two-cotyledon, bipolar, heart-shaped embryos, and no aberrant teratoma-like structures. Heart-shaped embryos developed and germinated normally on Murashige and Tucker basal medium with 0.5% sucrose and 1 μM gibberellic acid. Culture of embryogenic callus directly onto cellulose membranes also resulted in the development of normal heart-shaped embryos, indicating that glycerol induction of globular-stage embryos is not necessary. Heart-shaped embryos were not observed when the osmotic potential of the medium was increased by the addition of 2.5-15% polyethylene glycol; neither were they observed when the matric potential of the medium was increased by increasing the gelling agent concentrations of agar and Gelrite from 0.8% to 3% and 0.15% to 0.9%, respectively.

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