Germination and storability of calcium-alginate coated somatic embryos of mango ginger (Curcuma amada Roxb.)

Abstract

We developed a protocol to produce synthetic seeds of mango ginger (Curcuma amada Roxb.) for large-scale propagation, storage, and germplasm exchange. The seeds were produced by encapsulating somatic embryos in a calcium–alginate matrix. Embryogenic callus was induced from leaf sheath explants on 0.8% agar solidified with full-strength Murashige and Skoog (MS) medium supplemented with 3% (w/v) sucrose, 2.0 mg·L-1 2,4-dichlorophenoxyacetic acid (2,4-D) and 0.5 mg·L-1 6-benzyladenine (BA). Somatic embryos were induced by transferring the calli to 1/2 MS liquid medium supplemented with 3% (w/v) sucrose and 0.3 mg·L-1 BA. A synthetic seed coat for the somatic embryos that is uniform in size and shape was produced by incubating the embryos in 3% (w/v) sodium alginate in 1/2 MS liquid medium and exposure to 100 mM calcium chloride (CaCl2.2H2O) for 15 min. The highest germination percentage of synthetic seeds (91.66%) was achieved on 1/2 MS solid medium supplemented with 3% (w/v) sucrose and 0.25 mg·L-1 gibberellic acid (GA3) in darkness (24 h) for 35 days. Sucrose-dehydrated synthetic seeds showed better storability than fresh seeds. Synthetic seeds that were dehydrated in 8.55% (w/v) sucrose solution and stored at 4?C showed germination percentages of 88.10% after 30 days of storage and 54.16% after 120 days of storage. Plantlets were successfully acclimatized to ex vitro conditions (survival rate, 82.66%) and showed normal phenotypes. In a random amplified polymorphic DNA (RAPD) analysis, a monomorphic banding profile was obtained from the plants derived from synthetic seeds that had been stored at 4°C for 120 days. This confirmed the genetic fidelity of the plants derived from the stored artificial seeds.