Abstract: In order to maintain a high regeneration rate of plant callus with a long time, and to identify somaclnal variation, in this study, the embryonic calli of Zoysia matrella subcultured for 15 years were treated with different concentrations of epibrassinolide (EBL). The effects of BR the its growth and regeneration of long-term in vitro cultured callus were studied, and the optimal EBL concentration was selected to improve the growth and regeneration efficiency, which provided a reference for optimizing the subculture and regeneration system of Z. matrella. Meanwhile, to investigate the mechanism of the antioxidant system during embryogenesis of Z. matrella, catalase (CAT), peroxidase (POD), superoxide dismutase (SOD) activities, and malondialdehyde (MDA) content in callus and regenerated plants treated with EBL were examined. The results showed that 0.02 μmol·L^-1 EBL had the most significant promotion for callus growth and regeneration. The embryogenesis of callus was best maintained under the treatment of 0.02 μmol·L^-1 EBL, the diameter increased by 131.11% while the fresh weight increased by 43.8%. High concentrations of EBL (>0.10 μmol·L^-1) inhibited the embryogenic maintenance and regeneration of callus. The antioxidant reaction system in plants was activated by EBL treatment. The activity of POD decreased during subculture; the activities of CAT, POD, and SOD were improved during the regeneration. Additionally, the content of MDA decreased noticeably during the subculture and regeneration, which suggests that BR may regulate the physiological and biochemical processes during somatic embryogenesis by activating the antioxidant response and strengthening the antioxidant reaction in the process of subculture and regeneration. This study provides a solution to alleriated the problem that the regeneration rate of callus in long term culture decreases with the growth of culture time.

Key words: Zoysia matrella, brassinosteroid, callus growth, plant regeneration, antioxidant enzymes

摘要： 为了维持长期培养植物愈伤组组织的高再生率,以应用于体细胞无性系进行变异筛选。本研究以继代15年的沟叶结缕草[Zoysia matrella (L.) Merr.]胚性愈伤组织为材料,用不同浓度的表油菜素内酯(EBL)处理,研究油菜素甾醇(BR)对其生长和再生的影响,选出可提高长期离体培养愈伤组织生长及再生效率的最优EBL浓度。同时,测定EBL处理的愈伤组织和再生植株中过氧化氢酶(CAT)、过氧化物酶(POD)、超氧岐化酶(SOD)活性和丙二醛(MDA)含量等生理指标变化,探究BR调控沟叶结缕草体细胞胚发生过程中抗氧化系统的作用机理。结果表明,0.02 μmol·L^-1EBL对愈伤组织生长和再生的促进作用最明显,直径增长率和再生率分别为131.11%和43.8%,且愈伤组织胚性维持最佳。高浓度的EBL(>0.10 μmol·L^-1)抑制愈伤组织的胚性维持和再生。EBL处理激活了植物体内的抗氧化反应系统,继代生长中POD活性降低;再生过程中,CAT、POD、SOD活性均增强;继代和再生过程中MDA含量均降低,表明BR途径能激活愈伤组织的抗氧化系统的响应,加强继代和再生过程中的抗氧化反应,从而调控体细胞胚胎发生过程中的生理生化进程。本研究为缓解长期培养的愈伤组织再生率随着培养时间增长而下降的难题提供了解决方案。

关键词: 沟叶结缕草, 表油菜素内酯, 愈伤组织生长, 植物再生, 抗氧化酶