小菊悬浮细胞培养与植株再生研究

Studies on Cell Suspension Culture and Plant Regeneration of Dendranthema×grandiflorum with Small Inflorescences

以小菊‘七月红’品种为试材，进行了细胞悬浮培养及植株再生诱导的探讨。结果表明，MS＋1．0mg·L^-1 2，4-D＋0．2mg·L^-1 6-BA培养基适合从试管苗茎段有效诱导生长迅速、质地疏松的愈伤组织。获得的愈伤组织在MS＋0．5mg·L^-1 2，4-D＋0．2mg·L^-1 6-BA液体培养基中振荡培养，建立细胞悬浮培养体系。悬浮细胞的生长曲线呈上升的半抛物线型，细胞生长大致分缓慢生长期（0～2d）、对数生长期（2～10d）和停滞期（10d以后）；随着悬浮细胞生长，培养液pH值迅速下降。悬浮细胞固液双层培养表明，在培养基MS＋0．2mg·L^-1 KT＋0．2mg·L^-1 2，4-D上植板率最高；4℃低温2h处理能促进悬浮细胞生长，提高植板率；随着继代次数增加植板率呈下降趋势。培养1个月后，形成了直径约0．2cm的愈伤组织，将其转到MS＋2mg·L^-1 6-BA＋0．1mg·L^-1 NAA培养基后，继代4次，分化后出芽；分化芽在培养基MS＋0．5mg·L^-1 NAA上诱导生根，形成小植株。

Chrysanthemum ‘ Qiyuehong＇ with small inflorescence was employed for study of cell suspension and plant regeneration. The results showed that MS （Murashige and Skoog） medium with supplement of 1.0 mg · L^-1 2,4-dichlorophenoxyacetic acid （2,4-D） and 0. 2 mg · L^-1 6-benzylamino purine （6-BA） could efficiently induce rapid-growing and loose calli. Obtained calli were subsequently cuhured in liquid MS medium with 0. 5 mg ·L^-1 2,4-D and 0. 2 mg · L^-1 6-BA for establishing cell suspension system. The growth of suspension cuhured cells showed a parabola curve, mainly characterized by lagged growth phase （0 -2 d）, logarithmic growth phase （2 -10 d） and stationary growth phase （10 d later）, respectively. In addition, pH value of medium decreased rapidly during the cuhure process. Highest plant efficiency was obtained on MS medium with 0. 2 mg · L^-1 kinetin （KT） and 0. 2 mg· L^-1 2,4-D. Low temperature treatment at 4% for 2 hours could enhance the growth of cell, and increase the planting efficiency as well. However, the plant efficiency decreased with increasing of subcuhure times. Calli with a diameter of 0. 2 cm were obtained in one month. When they were transferred to MS medium containing 2. 0 mg ·L^-1 6-BA and 0. 1 mg · L^-1 naphthaleneacetic （NAA） and subcuhured for 4 times, shoots differentiated from the calli. Rooting plantlets were induced on MS with 0. 5 mg· L^-1 NAA.