ScienceAsia 30 (2004): 293-299 |doi: 10.2306/scienceasia1513-1874.2004.30.293

Cryopreservation of Shoot Tips of Dendrobium Walter Oumae by Encapsulation/Dehydration

Wanna Lurswijidjarus^a and Kanchit Thammasiri^b*

ABSTRACT: In vitro shoot tips of Dendrobium Walter Oumae were cryopreserved using the encapsulation/ dehydration technique. Shoot tips were encapsulated in calcium-alginate before preculture on modified Vacin and Went (1949) agar medium supplemented with 0.3, 0.5 and 0.7 M sucrose for 2 d at 252 ^oC, 37 mol m^-2s^-1 for 16 h per d. Encapsulated shoot tips were then dehydrated by incubation in the sterile air flow of a laminar air-flow cabinet for 0-10 h and immediately plunged into liquid nitrogen (LN). After recovering from LN and rapid thawing in a waterbath (402 ^oC), the survival ratio and regrowth were measured by a 2,3,5-triphenyl tetrazolium chloride (TTC) assay and a regrowth culture test, respectively. The highest survival ratio (16.18 mg living cells/100 mg total cells) and regrowth (13.33%) were obtained from encapsulated shoot tips that were previously precultured on 0.3 M sucrose agar medium for 2 d and sufficiently dehydrated for 6-8 h before storing in LN. The regrowth of plantlets was measured from length of shoot tips, number of shoots and roots which did not show any significant difference when air-dehydration and LN were tested. Finally, the regrown shoot tips could develop directly into complete plantlets without protocorms formation and they had normal morphology.

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a Department of Biotechnology, Faculty of Science, Mahidol University, Bangkok 10400, Thailand.
b Department of Plant Science, Faculty of Science and Institute of Science and Technology for Research and Development, Mahidol University, Bangkok 10400, Thailand.

* Corresponding author, E-mail: scktr@mahidol.ac.th

Received 21 Jul 2003, Accepted 3 Aug 2004