Effects of Growth Regulators and Temporary Immersion Bioreactor System on Shoot Multiplication and Root Induction of Anoectochilus burmanicus Cultured In Vitro
Keywords:
Anoectochilus burmanicus, in vitro propagation, auxin, cytokinin, temporary immersion bioreactorAbstract
Anoectochilus burmanicus is the terrestrial orchid with uniquely beautiful leaves and has been used for medicinal purposes. Nowadays, more and more A. burmanicus plants are being taken out of the forest. Therefore, there is a high risk of extinction. In this research, an effective method for micropropagation of A. burmanicus was developed. For shoot multiplication, single node explants were cultured on semi-solid ½ MS medium supplemented with BAP or TDZ (0.25, 0.50 and 1.00 mg/L) for 12 weeks. It was showed that adding 0.50 mg/L TDZ resulting in the highest average number of shoots at 3.1 shoots per explants. In addition, shoot multiplication was compared in semi-solid medium and the twin-flasks temporary immersion bioreactor (TIB) by testing feeding every 6 and 12 h for 5 and 10 min each time. After 12 weeks of cultivation, it was found that the TIB system could increase number of shoots and shoot length more than semi-solid medium. By feeding liquid medium every 12 h for 5 min each time, this gave the highest average number of shoots and average shoot length at 4.8 shoots per explants and 4.73 cm, respectively. For in vitro rooting, shoot explants were cultured with ½ MS medium without auxin or with 0.25, 0.50 and 1.00 mg/L IAA, and grown on semi-solid medium or in TIB system for 12 weeks. Result shown that cultivation in TIB system led to more growth in various aspects than the semi-solid medium. Plants grown in TIB system with medium containing 1.00 mg/L IAA showed the highest number of roots, root length, shoot height, number of leaves, leaf area and shoot fresh weight at 3.1 roots per shoot, 5.70 cm, 5.58 cm, 2.9 leaves per shoot, 595.80 mm², 1384.78 mg and 99.89 mg, respectively. After that, in vitro rooted plants were transplanted and acclimatized in clear plastic cups filled with peat moss for 4 weeks. It was found that all treatments had high survival rates and were not statistically different, with an average of 80-100%.
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