Sterilisation of Hyponex Medium by Chemicals without Autoclaving and Growth of Phalaenopsis Protocorms

For sterilization of Phalaenopsis culture medium without autoclaving, selected single sterilizing agents and in combinations were added to a 25ml Hyponex medium in a 120ml glass container. Treated liquid and solid media, supplemented with sterilizing agents, were compared to a control medium, autoclaved at 121°C for 15min. It was found that 90(L of 10% povidone-iodine, 150(L of 5.25% sodium hypochlorite, 150(L of 2% mercurochrome, 90(L of 2.5% iodine + 2.5% potassium iodine in combination with 10% providone-iodine (1:3) and 30(L of 2.5% iodine + 2.5% potassium iodide in combination with 2% mercurochrome showed 100% sterile conditions in liquid medium but provided 75, 100, 50, 75 and 80% sterile conditions, respectively, in solid medium. For growth of Phalaenopsis protocorms, 90(L of 10% povidone-iodine in liquid Hyponex medium gave the comparable growth of protocorms to control medium while 150(L of 5.25% sodium hypochlorite in solid medium provided the promising growth of protocorms. Growth of protocorms, whole fresh weight, numbers of leaf and root, root length and number of protocorm-like bodies, was discussed.

• Chockpisit Thepsithar
• Aree Thongpukdee

• Phalaenopsis
• sterilizing agents
• Hyponex medium
• sterile medium without autoclaving.

[1] A. D. Russell, "Similarities and differences in the responses of
microorganisms to biocides." Journal of Antimicrobial
Chemotherapy, vol. 52, no. 5, pp. 750-763, 2003.
[2] United State Patent 5750402. Compositions and methods to prevent
microbial contamination of plant tissue culture media. (Online).
Available; http://www. freepatentonline. com/ 5750402.html.
[3] U. Habiba, S. Reza, M. L. Saha, M. R. Khan, and S. Hadiuzzaman,
"Endogenous bacterial contamination during in vitro culture of table
banana: Identification and prevention." Plant Tissue Culture, vol. 12,
pp. 117-124, 2002.
[4] S. L. Teixeira, "The development of a new protocol that uses sodium
hypochlorite to replace the autoclaving procedure for establishing
axenic in vitro banana (Musa sp.) culture." Agricell Report, vol. 47,
no. 3, pp. 17-18, 2006.
[5] T. Yanagawa, M. Nagai, T. Ogino, and M. Maeguchi, "Application
of disinfection to orchid seeds, plantlet and media as a means to
prevent in vitro contamination." Lindleyana, vol. 10, no. 1, pp. 33-
36, 2006.
[6] M. Chansean, and I. Syoichi, "Conservation of Wild Orchids in
Cambodia by Simple Aseptic Culture Method." Proceedings of
NIOC, pp. 13-19, April, 2007.
[7] G. Nishimura, "Japanese orchids." In: Orchid Biology: reviews and
Perspectives II. Orchid seed germination and seedling culture - a
manual. Arditti, J. (Ed.). Ithaca: Cornell University Press, New York,
pp. 331-346, 1982.
[8] D. B. Duncan, "Multiple range and multiple F test." Biometrics, vol.
11, pp. 1-42, 1955.