(64(3):177-188)In Vitro Seed Germination and Micropropagation of Rehmannia glutinosa Libosch

Abstract

本研究利用地黃 (Rehmannia glutinosa Libosch) 種子進行無菌播種，地黃種子以0.6% 次氯酸鈉溶液消毒15 min 可獲得良好之殺菌效果，可明顯降低汙染率，並有最高發芽率，分別為5% 與80%。以無菌播種地黃小苗之葉片與葉柄作為微體繁殖材料，接種於含有3% 蔗糖、0.8% Bacto agar 及0.01 mg L-1 NAA 之MS 培養基 (以下簡稱基本培養基) 並添加0–2.0 mg L-1 BA 不同濃度，分別於光照與黑暗環境下培養。結果顯示，暗處理下培植體有較高之成活率，且葉柄培植體較葉片培植體之再生反應為佳，葉柄培植體於含有2.0 mg L-1 BA培養基中於黑暗環境下培養具有最高之存活率62%，可同時誘導出芽體與癒傷組織，分別為10% 與62%。另以地黃組培苗之頂芽莖段與莖中段2 種不同莖段部位作為微體繁殖材料，接種於添加不同蔗糖濃度 (3–9%)之基本培養基中培養，結果顯示以3% 蔗糖為最佳，成活率可達100%，9% 高蔗糖濃度為最差，會導致培植體褐化死亡。進一步利用組培苗之頂芽莖段、莖中段與近基部莖段3 種不同莖段部位培植體培養於含有3%蔗糖濃度並添加0–2.0 mg L-1 BA 濃度之基本培養基中誘導芽體，3 種培植體中以頂芽莖段表現最佳，成活率為80–100%，BA 濃度於0.5–1.0 mg L-1 之間可誘導出最多芽體，每1 培植體平均可誘導出27 個芽。莖段培植體與BA 濃度二者皆顯著影響地黃芽體之誘導率與培養存活率，且二者間具有交感效應。 In vitro studieswere conducted to establish protocol for micropropagation of Rehmannia glutinosa Libosch, an important Chinese herbal medicine. Results showed that seeds disinfection with 0.6% sodium hypochlorite for 15 min significantly reduced the contamination rate to 5% and obtained the highest germination rate to 80%. Comparing the regeneration rate of leaf and petiole explants from in vitro grown seedlings revealed that petiole explants growing on MS medium containing 0.01 mg L-1 1-Naphthaleneacetic acid with 2.0 mg L-1 Benzyladenine (BA) and cultivated in darkness obtained a better survival rate (62%) along with 10% shoot induction rate and 62% callus induction rate. Cultivating shoot tip and nodal stem segment of in vitro grow seedlings on the basal medium containing various sucrose concentrations (3–9%) showed that addition of 3% sucrose had the highest survival rate (100%). Furthermore, shoot tip, inter-nodal and basal nodal stem segments of seedlings were cultured on the basal medium containing 3% sucrose and various BA concentrations (0.0–1.0 mg L-1) to evaluate their regenerative potential. The results showed that shoot tip had the highest survival rate (80–100%) and had the most adventitious shoots induced per explant when cultured on the medium containing 0.5 mg L-1 BA. The results show that type of stem segment explants and BA concentration are key factors influence the micorpropagation of Rehmannia glutinosa Libosch