Influence Of Flower Inducing Of Dioscorea Zingberensis C.H.Wright In Vitro

Dioscorea zingberensis C.H. Wright, a native medicinal plant species in China, has a long history in application of medical materials. The highest content of diosgenin was reported to be 16.15 %. From 1970s up to now, many progresses have been made in breeding, distilling techniques of diosgenin and rapid propagation. In our study, we established a stable system of inducing inflorescences in vitro, which provided a new knowledge about flower formation and further opportunity for breeding. The main results are as follows: 1. Factors of inducing inflorescences in vitro1) Explants. In order to compare the induce difference due to explants, mature inflorescences, budds, immature inflorescences, shoot with immature inflorescences and shoot without inflorescences were sampled from strain No.4-73. The rates of explants on inducing inflorescences in vitro in different explants were calculated. The inducing rate from mature inflorescences was the highest (71.67 %), followed by the immature inflorescences (58.34 %), the axillary bud (45.00 %), and then the shoot with immature inflorescences(30 %). None of inflorescences was induced from the vegetative shoot in vitro.2) Origin of the explant. In order to compare the inducing capacity of different population of D. zingberensis, 5 populations, including No.323, No.4-73, No.5-88, No. 185 and No.31 were chosen as donor plant. Mature inflorescences from these 5 populations were excised as explant and inoculated in same medium. The result indicated that inducing efficiency has great variance in population. No.323 showed the highest rate (90.00 %) on inducing inflorescences in vitro, and No.30 was the lowest (33.33 %).3) Medium. No.323 was chosen as experimental material to research the Factors influencing of flower inducing in vitro, that were hormone, concentration of Fe²⁺-EDTA and sucrose, pH value.â‘ Hormone. When only one hormone was used, BA had the ability on inducing inflorescences in vitro, the highest rate (88.67 %) was obtained when 2.0 mg/L of BA was added into the medium,, KT, IAA, NAA, IBA, 2,4-D did not have the ability to induce inflorescences in vitro. Combination of BA with IAA, IBA, NAA had no effect to promote inflorescence induce, but it could improve the growth condition of the regenerated inflorescences. The best combination of hormone was BA(2.0 mg/L) and NAA(0.2 mg/L). Supplement of GA depressed the rate of inflorescence formation but put ahead the anthesis and prolonged the node length of the rachides.â‘¡Fe²⁺-EDTA. When the concentration of Fe²⁺-EDTA was 1/4 MS-Fe²⁺, 1/2MS-Fe²⁺, 1 MS-Fe²⁺, 2 MS-Fe²⁺. 4 MS-Fe²⁺, the rate was 86.67 %, 83.33 %,83.33 %, 50.00 %, 43.33 %. Lower concentration of Fe²⁺-EDTA increase the rateof flower formation while the higher concentration promoted the vegetativegrowth.â‘¢Sucrose. The rate of inducing inflorescences in vitro reduced when theconcentration of sucrose was improved. The appropriate concentration of sucrosewas 1.5 %-4.5 %.â‘£pH value. The rate of inducing inflorescences in vitro reduced when the pH valuewas improved. The optimum pH value was 5.8-6.0 .2. Subculture of regenerated inflorescencesDense callus was produced when explants, mature inflorescences, budds, immature inflorescences and shoot with immature inflorescences were induced in vitro. Some of the dense callus only contained inflorescences, and others contained inflorescences and vegetative branches. Inflorescences and vegetative branches were excised from the dense callus and then the dense callus and excised inflorescences were subcultured into new medium. The result showed that the dense callus would no longer regenerate new inflorescences, but the subcultured inflorescences maintained the capacity of florescence inducement as well as that from the inflorescences in vivo.3. Development of inflorescences formation in vitro1) The explants were sampled when cultured 0 d, 1 d, 3 d, 5 d, 7 d, 14 d, 25 d, 30 d. Development of the inflorescences formation was inspected with method of Paraffin Section. The inflorescence formed in vitro were initiated from the surface cells of the bract and perianth. The position of bud contacted with rachis extended when the explants had been cultured about 7 days. Small tubers formed at the extended position when the explants had been cultured about 20 days. Dense callus was produced when the explants had been cultured about 30 days, and then Multiple inflorescences were formed from the dense callus. Most of small flowers were flowering when the explants had been cultured about 100 days.2) We observed the process of regenerated male flower's formation and growth. Regenerated male flower has four anthers. Tetrads form through the meiosis of mother cells of microspore. At last, mature two nucleus pollens form by mother cells of microspore. So the process is the same as it grow in vivo.