Surface Sterilization Using Calcium Hypochlorite On Coconut (Xiem Red Dwarf) Zygotic Embryos

Abstract

Xiem Red Dwarf is considered as one of the new and important coconut varieties, which offers many socio-economic and environmental benefits in Vietnam. Embryo culture technique is now used to propagate and conserve this variety. Surface sterilization is one of most crucial parts in plant tissue culture. Sodium hypochlorite is used as a conventional disinfectant coupled with ethanol in various research; however, calcium hypochlorite is exerted in the same way in other researches. Calcium hypochlorite has been applied in different fields such as microbiology, cell reprogramming, and so on due to its long storage life and low risk for tissue culturists. In this study, zygotic embryos isolated from the mature Xiem Red Dwarf fruits (10 to 12 months post pollination) are used as plant materials and sterilized by 0.5% (w/v) of sodium hypochlorite and 0.5% (w/v) of calcium hypochlorite with 70%(v/v) of ethanol. Data showed that the effect of calcium hypochlorite is as good as sodium hypochlorite in terms of surface sterilization and germination rate of coconut zygotic embryo. This result suggests that calcium hypochlorite could be used as an alternative disinfectant in the surface sterilization protocol of coconut embryo. In addition, data indicated that ethanol has the negative effect on the subsequent growth of coconut plantlets due to its oxidative and inhibitory characteristics that leads to a decrease in shoot length and root length. So it is recommended that ethanol should be removed from surface sterilization protocol for coconut embryo culture. The final experiment assessing effect of different concentrations of calcium hypochlorite on the growth of in vitro seedling showed that 0.5% of calcium hypochlorite is the optimal concentration. Higher concentrations of calcium hypochlorite (3 and 6%) led to a signigicant reduction in the seedling growth. In summary, it is evident that calcium hypochlorite can be effectively used to aseptically prepare coconut embryo explant and it can also support in vitro growth of seedlings. This study also suggests that ethanol should be excluded from the surface sterilization protocol used for coconut embryo explant.