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Determination of Appropriate Conditions for Medium-term in Vitro Storage of Coconut (Cocos nucifera L.) embryos.
Thesis Abstract:
Experiments were conducted to determine the best conditions for in vitro storage of zygotic embryo of coconut. To understand cytological changes that might have occurred during the in vitro storage, morpho-anatomical observations on slide preparations of stored and unstored embryos were also done.
Mature embryos of tall coconut variety “Tagnanan” from PCA Zamboanga Research Center were cultured in MS media with and without mannitol (0.01, 0.2, 0.3, and 0.4M) under three storage temperatures (5, 15, and 25°C) and three light conditions (dark, 3 W.m-2, 6 W.m-2). Mannitol, temperature, and light treatments were combined with half and full strength basal media. Responses of the embryos to the different culture conditions were determined at three, six, and nine months of culture.
The results indicated that embryos could be kept in the MS medium for three months without significant loss in viability. Prolonging the storage to six months led to 60 percent loss of viability. Storage for nine months in full-strength medium rendered all embryos nonviable while storage in half-strength medium left 15 percent of the embryos still able to germinate.
Addition of mannitol up to 0.3M did not have any significant effect on embryo viability during storage or on their subsequent germination after storage. However, seedlings derived from embryos kept in media with 0.3M mannitol showed some morphological abnormalities such as formation of root without shoot, shoot without root and stunted seedlings. Higher level of mannitol (0.4M) caused death of the embryos, even for the short (three months) storage period.
Storage temperatures below 25°C significantly decreased the size of embryo and germination rate. Storage at 5°C resulted in complete loss of viability by the third month. The 25°C storage temperature proved optimum particularly when combined with half-strength nutrient medium. This treatment gave the highest percentage (90%) of germination of embryo and produced normal seedlings.
Highest percent germination (90%) was observed with embryos stored under high light intensity (6 W.m-2). However, lower percentage of morphologically normal seedlings was obtained from embryos stored under this light intensity than from embryos stored in the dark or a low light intensity (3 W.m-2).
The possibility of preserving coconut embryos in vitro offered a complementary method to field gene banking. Overall, the results of study indicated the need for further experimentation on the culture media and culture environment to determine the best conditions for medium-term storage of coconut embryos. The technique still has potential use for short-term storage and for in vitro collection, exchange and distribution of coconut germplasm.