| Plant latin name | Glycyrrhiza glabra Linne |
| Literature code | Glycyrrhiza_glabra-Ref-1 |
| Reference | Kohjyouma M et al., Plant Tissue Culture Letters 12: 145-149 (1995) |
| Summary | Stem segments with axillary buds of Glycyrrhiza glabra L. were cultured on modified MS medium supplemented with a-naphthalene acetic acid (NAA) and 6-benzylaminopurine (BAP). Formation of multiple shoots was obtained on the medium provided with 1.00 mg/l BAP after 40 days of culture. Each shoot was cut and transplanted onto the medium supplemented with NAA and BAP. After 80 days of culture, the rooting of shoots was well established in the media containing 0.01-0.50 mg/l NAA and these shoots subsequently developed into whole plants. |
| Objectives | Establishment of a rapid in vitro propagation method for Glycyrrhiza glabra |
| Materials | Glycyrrhiza glabra L. plant cultivated at Kumano Medicinal Plants Garden of Saraya Co., Ltd. |
| Explant | The subterranean stems of Glycyrrhiza glabra were cut into the 15-20 cm lengths. They were washed with tap water for 1-2hr and soaked in 5% calcium hypochlorite solution for 10 min. They were transplanted to vermiculite, and cultivated in a growth chamber at 25 ± 2℃ in continuous light of fluorescent lamps (about 3,000 lux). After one month of cultivation, the 15-20 cm shoots growing in the pots were
divided into 5 cm long segments with a few axillary buds. |
| Initial culture | Shoots (15-20 cm in length) were washed with tap water for 1 hr, surface sterilized with 1% sodium hypochlorite solution with a drop of Tween 20 (1 drop/50 ml) for 5 min., and then rinsed with sterile water 3 times. They were cut into 1 cm long segments each
containing one axillary bud using sterilized scissors. The sterilized segments were cultured in 16 kinds of modified Murashige and Skoog media which organic components were changed to one of Gamborg B5 medium, each containing both NAA at concentrations 0, 0.04, 0.20, l.00 mg/l and BAP at concentrations 0, 0.2, 1.0，5.0 mg/l. All basal
media were provided with 3% sucrose and solidified with 0.8% agar. They were statically cultured in bottles (180 ml) containing about 30 ml medium adjusted to pH 5.5 before autoclaving. The culture bottles were covered with alminium sheets and exposed to light for 16 hr (3, 000 lux at 25±2℃) .
After 40 days of culture, morphogenetic changes were observed. The media containing BAP without NAA were effective in the formation of multiple shoots. These shoots were abnormally shaped compared to the mother plant. Calli formation was mainly observed on the media provided with 5.0 mg/l BAP with 0.04-0.20 mg/l NAA or without NAA. |
| Shoot multiplication | The effect of BAP on shoot formation was tested with adding various concentrations of BAP (0.25, 0.50, 1.00, 2.00 mg/l) to the basal medium. 50 days after culture initiation, the number of shoots (more than 1 cm in length) per explant was recorded for each culture. The highest number of shoots, 4. 6 per explant, was obtained on the medium with 1.00 mg/l BAP. Therefore this concentration of BAP was selected as a best condition for the formation of multiple shoots for use in the following experiment. |
| Rooting | After 40 days of primary culture, the newly-formed multiple shoots were cut into individual shoots, and transferred onto fresh rooting media supplemented with various combinations of NAA (0.05, 0.10, 0.50 mg/l) and BAP (0, 0.01 mg/l). When the shoots were cultured in the rooting media for 40 days, they were transferred onto the same type of fresh media (50 ml) solidified with
0.6% agar in Erlenmeyer flasks (500 ml). When the newly formed shoots were transferred to the rooting media for 80 days, root initiation
and shoot elongation were observed. The highest frequency of root formation was obtained on the medium with 0.05-0.10 mg/l NAA. In this condition, development of the shoots and length and number of leaves were as in a normal plants. The addition of BAP did
not increase root formation. |
| Acclimation | The plantlets developed on the rooting medium were rinsed with water and transplanted into pots
with vermiculite and covered with a plastic film to maintain high humidity. |
| Planting | The plantlets were grown in a greenhouse. |
| Cultivation conditions | |
| Traints of regenerants | |
| Ingredients analyzed | |
| Extraction | |
| Analitical methods | |
| Notes | |