Cryopreservation of immature seeds of Bletilla striata by vitrification

An efficient protocol was established for the cryopreservation of immature seeds of a terrestrial orchid, Bletilla striata. Immature seeds collected 2-4 months after pollination (MAP) were treated using three different cryogenic procedures: (1) direct plunging into liquid nitrogen, (2) vitrification, and (3) vitrification with preculture. When immature seeds collected 3 MAP and 4 MAP were precultured for 3 days on New Dogashima medium supplemented with 0.3 M sucrose and cryopreserved by vitrification, the survival rate after preservation, as assessed by staining with 2,3,5-triphenyltetrazolium chloride, was 92% and 81%, respectively. Immature seeds thus treated showed no decrease in germination rate relative to untreated immature seeds, and they developed into normal plantlets in vitro.