Cryopreservation of zygotic embryos of a Japanese terrestrial orchid (Bletilla striata) by vitrification

• Published in: Plant cell reports 1997-09, Vol.16 (11), p.754-757
• Main Authors: ISHIKAWA, K, HARATA, K, MII, M, SAKAI, A, YOSHIMATSU, K, SHIMOMURA, K
• Format: Article
• Language: English
• Subjects: Biological and medical sciences; Biotechnology; Embryos; Eukaryotic cell cultures; Fundamental and applied biological sciences. Psychology; In vitro embryo culture; Methods. Procedures. Technologies; Plant cells and fungal cells; Sucrose
• ISSN: 0721-7714; 1432-203X
• DOI: 10.1007/s002990050314

The seeds of a Japanese terrestrial orchid (Bletilla striata Rchb.f.) were germinated and cultured on solidified new Dogashima (ND) medium for 10 days. These embryos were then precultured on ND medium supplemented with 0.3 M sucrose for 3 days at 25°C in continuous dark. The embryos were then overlaid with a mixture of 2 M glycerol and 0.4 M sucrose for 15 min at 25°C and finally dehydrated with highly concentrated vitrification solution (PVS2) for 3 h at 0°C prior to immersion into liquid nitrogen for 30 min. After rapid warming, the embryos were washed with liquid ND medium supplemented with 1.2 M sucrose for 20 min and then plated on ND medium. Successfully vitrified and warmed embryos developed into normal plantlets. The rate of plant regeneration amounted to about 60%. This vitrification method appears to be a promising technique for cryopreservation of orchids.