Abbasi Bilal Haider, Ali Huma, Yücesan Buhara, Saeed Sabahat, Rehman Khalid, Khan Mubarak Ali
Department of Biotechnology, Quaid-i-Azam University, Islamabad, 45320, Pakistan.
Department of Biotechnology, Bacha Khan University, Charsada, KP, Pakistan.
3 Biotech. 2016 Jun;6(1):71. doi: 10.1007/s13205-016-0366-1. Epub 2016 Feb 16.
In present report effects of explants type, basal media and plant growth regulators (PGRs) were tested for induction of indirect somatic embryogenesis in medicinally important plant Silybum marianum L. Leaf, petiole and root explants were exploited in vitro on B5 (Gamborg), SH (Schenk and Hildebrandt) and MS (Murashige and Skoog) media for induction of embryogenic callus followed by somatic embryogenesis. Highest callus induction frequency (76 ± 4.8 %) was recorded when petiole explants of in vitro derived plantlets were cultured on B5 medium supplemented with 1.5 mg l 2,4-dichlorophenoxyacetic acid (2,4_D) in combination with 1.5 mg l Thidiazuron (TDZ). Induction and multiplication of somatic embryos were observed, when the embryogenic calluses were sub-cultured on to B5 medium containing 0.5 mg l 2,4-D plus 1.5 mg l TDZ. At this PGRs treatment, 77 % of the cultures responded with 39.1 somatic embryos per callus. Furthermore, MS0 medium was indicated more reponsive for growth and maturation of somatic embryos. Analysis of biochemical markers during various growth phases in somatic embryogenesis revealed that somatic embryos exhibited highest level of total carbohydrate, starch, ascorbic acid and total free amino acids. However, higher protein levels were detected in non-embryogenic callus. Nevertheless, considerable amount of silymarin (4.1 mg g DW) was detected in somatic embryos than other growth phases. Thus, the present study concluded that biochemical and physiological changes during embryogenesis are influenced by interplay of explants type, basal media and PGRs.
在本报告中，对药用植物水飞蓟（Silybum marianum L.）外植体类型、基础培养基和植物生长调节剂（PGRs）在间接体细胞胚胎发生诱导方面的作用进行了测试。利用叶片、叶柄和根外植体在B5（甘博格）、SH（申克和希尔德布兰特）和MS（Murashige和Skoog）培养基上进行体外培养，以诱导胚性愈伤组织，随后进行体细胞胚胎发生。当将体外培养的幼苗的叶柄外植体接种在添加了1.5 mg/L 2,4-二氯苯氧乙酸（2,4-D）和1.5 mg/L噻二唑隆（TDZ）的B5培养基上时，愈伤组织诱导频率最高（76±4.8%）。当将胚性愈伤组织继代培养到含有0.5 mg/L 2,4-D和1.5 mg/L TDZ的B5培养基上时，观察到体细胞胚胎的诱导和增殖。在此植物生长调节剂处理下，77%的培养物有反应，每个愈伤组织产生39.1个体细胞胚胎。此外，MS0培养基对体细胞胚胎的生长和成熟表现出更高的响应性。对体细胞胚胎发生不同生长阶段的生化标记物分析表明，体细胞胚胎中总碳水化合物、淀粉、抗坏血酸和总游离氨基酸水平最高。然而，在非胚性愈伤组织中检测到较高的蛋白质水平。尽管如此，与其他生长阶段相比，在体细胞胚胎中检测到了相当数量的水飞蓟素（4.1 mg/g干重）。因此，本研究得出结论，胚胎发生过程中的生化和生理变化受外植体类型、基础培养基和植物生长调节剂相互作用的影响。
