Embryogenic cultures of Dioscorea bulbifera were cryopreserved using an encapsulation-dehydration procedure with subsequent plant regeneration. Embryogenesis was induced by culturing in vitro grown axillary bud meristems on MS medium supplemented with 2.0 mg l−1 2,4-D. After cryopreservation, recovery growth of embryogenic culture up to 53.3% was recorded when excised proliferating embryogenic cultures of 1.5-2.0 mm in diameter were: encapsulated in 3% calcium alginate containing 0.15 M sucrose followed by preculturing with 0.5 M sucrose for 3 d; dehydrated in the laminar air flow for 4 h, thereby reducing the bead moisture content to 19.4% (fresh weight basis); plunged into liquid nitrogen; thawed at 40°C; and cultured on recovery growth medium, i.e. MS supplemented with 2.0 mg l−1 2,4-D and 0.3 mg l−1 BAP. However, preculturing for an extended period of 7 d increased the recovery growth further to 67.8%. During recovery growth the embryogenic tissue protruded out of the beads without loss of structural integrity of the cryopreserved embryos. Subculturing of these cultures on to embryo conversion medium, i.e. MS medium with 0.5 mg l−1 zeatin and 400 mg l−1 glutamine, resulted in production of plantlets through embryo conversion. The regenerated plantlets exhibited the same morphology as that of originally maintained in vitro plantlets and were established in vivo, in a net house with 80% success.
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Document Type: Research Article
Publication date: 01 November 2009
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CryoLetters is a bimonthly international journal for low temperature sciences, including cryobiology, cryopreservation or vitrification of cells and tissues, chemical and physical aspects of freezing and drying, and studies involving ecology of cold environments, and cold adaptation
The journal publishes original research reports, authoritative reviews, technical developments and commissioned book reviews of studies of the effects produced by low temperatures on a wide variety of scientific and technical processes, or those involving low temperature techniques in the investigation of physical, chemical, biological and ecological problems.