Dhir Richa, Shekhawat G S, Alam Afroz
Department of Bioscience and Biotechnology, Banasthali University, Rajasthan, 304022, India.
Appl Biochem Biotechnol. 2014 Aug;173(8):2267-78. doi: 10.1007/s12010-014-1032-x. Epub 2014 Jun 29.
An improved procedure has been developed for efficient somatic embryogenesis in Anethum graveolens. Green friable embryogenic callus was obtained from hypocotyl segments on medium augmented with 2,4-dichlorophenoxyacetic acid (2,4-D). The highest embryogenic callus induction frequency of 87 % was obtained on Murashige and Skoog (MS) medium containing 1.13 μM 2,4-D. At lower concentration of 2,4-D (0.34 μM) callus turned dark in color and slow growing. Embryogenic cultures (76 %) responded with a mean number of 43 globular and 18 heart stage embryos. Somatic embryo maturation and subsequent conversion into plantlets took place on MS lacking growth regulators. Maximum number of somatic embryos developed on MS medium was 128.3 (per flask) and a plantlet conversion of 82 % was observed. Calcium alginate beads were produced by encapsulating somatic embryos. Highest percent germination (83 %) was observed on 0.8 % agar solidified MS medium with the plantlets acquiring an average length of 2.1 cm. Encapsulated somatic embryos could be stored at 4 °C up to 60 days with a conversion frequency of 49.3 %. Highest protein and proline content has been observed in embryogenic callus with small globular embryos. During morphological differentiation of the somatic embryos, changes in the antioxidant enzymatic system were observed. Superoxide dismutase (SOD) activity increased during initial stages and decreased catalase (CAT), peroxidase (POD), and ascorbate peroxidase (APX) activities were detected.
已开发出一种改进方法，用于在莳萝中高效诱导体细胞胚胎发生。在添加了2,4-二氯苯氧乙酸（2,4-D）的培养基上，从下胚轴切段获得了绿色易碎的胚性愈伤组织。在含有1.13 μM 2,4-D的Murashige和Skoog（MS）培养基上，胚性愈伤组织诱导频率最高可达87%。当2,4-D浓度较低（0.34 μM）时，愈伤组织颜色变深且生长缓慢。胚性培养物（76%）平均产生43个球形胚和18个心形胚。体细胞胚胎在不含生长调节剂的MS培养基上成熟并随后转化为小植株。MS培养基上发育的体细胞胚胎数量最多为128.3个（每瓶），小植株转化率为82%。通过包裹体细胞胚胎制备了海藻酸钙珠。在0.8%琼脂固化的MS培养基上观察到最高发芽率（83%），小植株平均长度为2.1 cm。包裹的体细胞胚胎可在4℃下保存60天，转化率为49.3%。在含有小球形胚的胚性愈伤组织中观察到最高的蛋白质和脯氨酸含量。在体细胞胚胎的形态分化过程中，观察到抗氧化酶系统的变化。超氧化物歧化酶（SOD）活性在初始阶段增加，而过氧化氢酶（CAT）、过氧化物酶（POD）和抗坏血酸过氧化物酶（APX）活性降低。
