HIGH FREQUENCY SEED GERMINATION AND SEEDLING GROWTH IN VITRO IN ACAMPE RIGIDA ORCHID

Abstract

The percent germination of Acampe rigida seeds in vitro more than doubled when half-strength MS (HMS) medium solidified with agar was supplemented with various organic extracts. The percent germination was highest at 91.0% when 200 ml L-1 of rice extract was added to the HMS medium. The addition of 200 ml L-1 of potato extract resulted in 86.1% germination and the addition of 200 ml L-1 of wheat extract resulted in 85.2% germination. Germination percentage on HMS medium without supplement was 40.5%. Final counts were taken 3 months after the seeds were sown in vitro. Germinated seeds generally develop into protocorms and then develop into mini-seedlings. When protocorms were sub-cultured, some of the protocorms continued to multiply, producing new protocorms, while others began to develop into mini-seedlings. The addition of certain supplements to the HMS medium increased the number of multiplying protocorms in the cultures. However, the addition of organic extracts of banana and potato combined with the addition of 2 g L-1 activated charcoal in HMS medium were effective for protocorm multiplication. Among other media tested as suitable subculture media, NP (New Phalaenopsis) medium supplemented with 30 g L-1 maltose was the best for protocorm multiplication. After 2 months in fresh culture on this NP medium, 0.05 g protocorm average fresh weight increased to an average of 5.397 g. In this culture, there was an average of 185±5.1 new protocorms and an average of 107±3.1 mini-seedlings produced. Subsequent growth of seedlings on fresh medium in vitro was also affected by different supplements added to HMS medium. Among carbohydrates trialed, the addition of 30 g L-1 sucrose enhanced seedling growth when compared to maltose or sorbitol as alternative carbon sources. The addition of rice extract at 200 ml L-1 along with 2.0 g L-1 charcoal was the best for shoot growth in vitro compared with other treatments. Healthy seedlings grown on organic extracts in vitro were acclimatized in vivo on tissue paper, cloth towels and brown cardboard for 3 weeks in a shade house. Among these treatments, the highest percentage of successful conversion (96.8%) was obtained with tissue paper whereas the lowest conversion rate was for seedlings placed on brown cardboard. Growth of seedlings on a fresh weight basis during acclimatization was also higher in tissue paper compared to the other substrates. A high frequency seedling production protocol for commercial use has been established based upon this experimental data.