参薯愈伤组织诱导及其悬浮细胞培养体系的建立  

Induction of Dioscorea alata Callus and Establishment of Its Suspension Cell Culture System

作　　者：梅朋飞 吴晓龙 庄赛伟 王颖[1] 黄小龙[1] 黄东益[2] 夏薇[2] 吴文嫱[1] Mei Pengfei;Wu Xiaolong;Zhuang Saiwei;Wang Ying;Huang Xiaolong;Huang Dongyi;Xia Wei;Wu Wenqiang(College of Life Science,Hainan University,Haikou,570228;College of Tropical Crops,Hainan University,Haikou,570228)

机构地区：[1]海南大学生命科学学院,海口570228 [2]海南大学热带作物学院,海口570228

出　　处：《分子植物育种》2024年第3期840-848,共9页Molecular Plant Breeding

基　　金：海南省自然科学基金项目(2019RC116);国家自然科学基金地区项目(32160470)共同资助。

摘　　要：为确定参薯愈伤组织诱导的最佳激素浓度及悬浮细胞培养条件,以3种基因型参薯(Dioscorea alata)组培苗的腋芽茎段为材料,采用三因素三水平正交实验来探究不同浓度组合的6-BA、NAA和毒莠定(picloram,PIC)对愈伤组织诱导率的影响;为了初步建立愈伤组织悬浮细胞培养体系,采用单因素实验探讨初始悬浮细胞接种量、蔗糖浓度、摇床转速、酪蛋白浓度和光照时间对其细胞生物量的影响。实验结果表明:在1.0 mg/L 6-BA+1.0 mg/L NAA+0.5 mg/L PIC激素浓度组合下,参薯Da70号组培苗(Da70)的愈伤诱导率为90%,愈伤状态较好,不同激素对愈伤诱导率的影响分别为NAA>PIC>6-BA。在单因素悬浮细胞培养实验中,10 mL的初始悬浮细胞接种量、10 g/L的蔗糖浓度、100 r/min的摇床转速、50 mg/L的酪蛋白浓度和0 h光照有利于细胞生物量的增加。参薯悬浮细胞生长总体上呈现“S”型,且在第18天时,鲜重为0.1593 g,干重为0.0423 g,细胞活力OD值为0.4850,达到峰值。本研究为后续原生质体的分离以及功能基因的亚细胞定位提供依据。In order to determine the better hormone concentration and suspension cell culture conditions induced by Dioscorea alata callus,the axillary bud stem segments of three genotypes of Dioscorea alata's tissue culture seedlings were used as materials.And a three-factor,three-level orthogonal experiment was used to investigate the effects of different combinations of 6-BA,NAA and PIC on the induction of healing tissues.In order to initially establish a healing tissue suspension cell culture system,single-factor experiments were used to explore the effects of initial suspension cell inoculum,sucrose concentration,shaker speed,casein concentration,and light time on their cell biomass.The experimental results showed that under the combination of 1.0 mg/L 6-BA+1.0 mg/L NAA+0.5 mg/L PIC hormone concentration,the healing induction rate of Da70 was 90% and the healing status was better,and the effects of different hormones on the healing induction rate were as follows:NAA>PIC>6-BA.In the single-factor suspension cell culture experiment,an initial suspension cell inoculum of 10 mL,a sucrose concentration of 10 g/L,a shaker speed of 100 r/min,a casein concentration of 50 mg/L,and 0 h of light facilitated the increase in cell biomass.The growth of the suspension cells of the crop showed an overall"S"shape and reached a peak of 0.1593 g fresh weight,0.0423 g dry weight and 0.4850 cell viability OD at the 18th day.The results of this study can provide the basis for the subsequent isolation of protoplasts and the subcellular localization of functional genes.

关 键 词：参薯 组培苗 愈伤组织 悬浮细胞 培养条件 

分 类 号：S632.1[农业科学—蔬菜学]