Efficient Differentiation of Adventitious Embryos from Cotyledon Culture of Camellia sinensis and Other Camellia Species

Abstract

Adventitious embryo formation from Camellia sinensis cotyledons was studied by tissue culture. The effects of benzyladenine (BA) concentration in culture medium, the explant size and the developmental stage of cotyledon used, and also interspecific and varietal differences were investigated in cotyledon culture of C. sinensis and other species of Camellia.
1) In cotyledon culture of C. sinensis cv. Yabukita, M&S medium supplemented with BA (1.0-5.0 mg/l) increased the rate of the differentiation to 30% or higher from 16% of that without the addition of BA. The differentiation of adventitious embryos in a few basal medium and a few combinations of either BA plus naphthalene acetic acid (NAA) or BA plus gibberellin A3 (GA) were of lower rates and less stable than that of BA (1.0-5.0 mg/l) alone.
2) Differentiation of adventitious embryos was remarkably influenced by the size of explants of cotyledon. The larger the explants of cotyledon were, the higher the differentiation rate of adventitious embryos occured.
3) The optimum timing of cotyledon culture for the highest differentiation rate of adventitious embryos was found to be late September to mid October (Stage 2) using ripe seeds. The rates of the differentiation were lowered when unripe seeds (Stage 1) or stored ripe seeds (5°C for 2-3 months, Stage 3) were used for the culture. Also, bacterial concentration of explants became more and more frequent as the storage periods of seeds became longer (Stage 3).
4) Remarkable interspecific and varietal differences were observed in the rates of the differentiation of adventitious embryos. The rates among C. sinensis ranged from zero to near fifty per cent : High rates were obtained with 'Yabukita', 'Kurasawa' while, these of 'Houryoku', 'Da-Yeh Oolong', 'Manipuri 9', 'Chi-Shin Dah-Pan' and 'Satumabeni' were less than 6%. On the other hand, the differentiation rate of adventitious embryos of other Camellia species were high in comparison with C. sinensis. Especially high differentiation rate of adventitious embryos were obtained with C. japonica (48-58%), C. sasanqua (59-81%) and C. brevistela (93%). Also the high differentiation rate of 69% was obtained with the cotyledon of interspecific hybrid between C. japonica and C. granthamiana.