Studies on single and double layered biocompatible encapsulation of somatic embryos in Albizia lebbeck and genetic homogeneity appraisal among synseed derived lines through ISSR markers
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Embryogenic synseeds were prepared in Albizia lebbeck by encapsulating cotyledon stage somatic embryos derived from in vitro maintained embryogenic cultures in different types of Ca-alginate beads. The germination rate of somatic embryos was affected significantly by the bead type, matrix composition and germination substrate. A matrix made of 3% Na2-alginate complexed with 100 mM CaCl2·2H2O for a hardening period of 20 min provided uniform encapsulation of somatic embryo. Among different types of synseeds, type IIA, wherein somatic embryos encapsulated in a single layer of Ca-alginate matrix composed of MS medium supplemented with 2 g L−1 activated charcoal and 1.0 µM gibberellic acid (GA3) as reconstituted endosperm, was found to be the most efficient type having maximum germination rates (88.6 ± 0.51%). Incorporation of GA3 in the alginate beads stimulated greater germination of somatic embryos as against GA3 supplementation in the germination substrate. Further, viability studies on short term cold (4 °C) storage of different types of embryogenic synseeds revealed that double layered synseeds (DLS) were found comparatively more robust to withstand longer storage durations than single layered synseeds as evident by greater germination rates of the former after 4–8 weeks of refrigerated storage. Also, the elevated levels of antioxidative enzymes (superoxide dismutase, catalase, ascorbate peroxidase and glutathione reductase) and leaf proline content in the plantlets derived from DLS reveals the possible role of alginate coatings in conferring alleviation to low temperature stress generated during different storage durations. Similar Inter simple sequence repeat profiles of embryogenic synseeds derived plantlets and mother tree nullifies the possible occurrence of somaclones, thereby establishing the efficacy of synseed technology for clonal propagation of A. lebbeck germplasm.
Encapsulation of cotyledonary stage somatic embryos combined with high in vitro germination rates provides a feasible tool for interim storage and exchange of A. lebbeck germplasm.
KeywordsAntioxidative enzymes Encapsulation Gibberellic acid ISSR Proline Somatic embryos
Double layered synseeds
Inter simple sequence repeats
Single layered synseeds
Taiba Saeed gratefully acknowledges UGC for providing the financial assistance under the scheme of Maulana Azad National Fellowship (File No. MANF-MUS-UTT-2624). Authors are also thankful to DST-SERB, for financial support in the form of Young Scientist Projects (Vide Nos. SB/FT/LS-364/2012 and SB/YS/LS-156/2013).
AS designed the research study and extended overall guidance during the experimentation. TS conducted the experimental work, analysed data and wrote the manuscript. SS helped TS in performing molecular experiment and drafting manuscript. All authors read and approved the final manuscript.
Compliance with ethical standards
Conflict of interest
Authors declare that they have no conflict of interest.
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