Studies on initial tissue culture techniques of mangrove tree species in Okinawa, Japan

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In this study, attempts have been made to develop initial tissue culture techniques of several mangrove tree species distributed in Okinawa, Japan. The samples were collected from trees of Bruguiera gymnorrhiza, Kandelia candel, Rhizophora stylosa (Rhizophoraceae) and Hibiscus tiliaceus (Malvaceae) in Okinawa island, and trees of Avicennia marina (Verbenaceae) and Sonneratia alba (Sonneratiaceae) in lriomote island. Materials used for inducing calli were viviparous seeds of B. gymnorrhiza, K. candel and R. stylosa, semi-viviparous seeds of A. marina, and seeds of H. tiliaceus and S. alba. In the callus culture, viviparous seeds were cut in disc shaped particles in 5 mm thickness, then put on the media. Two regents of phytohormones, Kinetin as a cytokinin and 2, 4-D (2, 4-dichlorophenoxyacetic acid) as a auxin, were added for inducing calli to MS medium (Murashige and Skoog medium, 1962). Few contaminations were observed for using a modified sterilisation method combined with the ethyl alcohol and sodium hypochlorite solutions method.There was no significant difference in the rates of callus inductions between any growing stages of viviparous seeds of B. gymnorrhiza, K. candel and R. stylosa when both kinetin and 2, 4-D were added to the callus inducing media. Calli were induced from hypocotyls of non-symbiotic seedlings of H. tiliaceus. The subcultures were done at the earlier stages of inducted calli of viviparous seeds, because those calli at the later stages became harder, and not grew well on the subculture media.