Cryopreservation of Bletilla striata Mature Seeds, 3-Day Germinating Seeds and Protocorms by Droplet-Vitrification

Authors: Jitsopakul, Nipawan; Thammasiri, Kanchit; Ishikawa, Keiko

Source: Cryoletters, Volume 29, Number 6, November 2008 , pp. 517-526(10)

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Abstract:

Droplet-vitrification was studied for the cryopreservation of Bletilla striata mature seeds (0 day after sowing), 3-day germinating seeds and protocorms (6, 9 and 12 days after sowing). Mature seeds, 3-day germinating seeds and 6-day old protocorms were precultured in liquid medium supplemented with 0.3 M sucrose for 3 h on a shaker (110 rpm) and then dehydrated with 2 M glycerol and 0.4 M sucrose in liquid medium (loading solution) for 15 min and exposed to PVS2 solution for 60 min at 25°C. The plant materials were then immersed in liquid nitrogen, rewarmed rapidly and cultured on solidified ND medium supplemented with 3% sucrose for recovery. After cryopreservation, the highest germination percentage of mature seeds, 3-day germinating seeds and survival of cryopreserved 6-day old protocorms was 93%, 91% and 84%, respectively. For 9-day old protocorms, highest survival (66%) after cryopreservation was achieved after preculture with 0.5 M sucrose for 3 h on a shaker, dehydration with loading solution for 15 min, exposure to PVS2 solution for 40 min at 25°C, and culture on solidified ND medium supplemented with 480 mg l^-1 ammonium nitrate and 3% sucrose. No survival was observed in cryopreserved 12-day old protocorms.

Keywords: BLETILLA STRIATA; CRYOPRESERVATION; SEEDS; PROTOCORMS; DROPLET-VITRIFICATION

Document Type: Research article

Publication date: 2008-11-01

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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.

Cryopreservation of Bletilla striata Mature Seeds, 3-Day Germinating Seeds and Protocorms by Droplet-Vitrification

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