Raji Mohammad Reza, Lotfi Mahmoud, Tohidfar Masoud, Zahedi Bahman, Carra Angela, Abbate Loredana, Carimi Francesco
Department of Horticulture, Faculty of Genetics and breeding of vegetables, University of Tehran, Pakdasht, Tehran, 3391653775, Iran.
Department of Plant Biotechnology, Faculty of Life Science and Biotechnology. G. C., Shahid Beheshti University, Tehran, Iran.
Protoplasma. 2018 May;255(3):873-883. doi: 10.1007/s00709-017-1194-9. Epub 2017 Dec 17.
A new protocol for in vitro regeneration through direct somatic embryogenesis for two muskmelon cultivars (Cucumis melo L., "Mashhadi" and "Eivanaki") is reported. Somatic embryos were obtained culturing 4- and 8-day-old cotyledons, seeds, and hypocotyls on Murashige and Skoog medium supplemented with three different hormonal combinations never tested so far for melon (naphthoxyacetic acid (NOA) + thidiazuron (TDZ), NOA + 6-banzylaminopurine (BAP), and 2,4-dichlorophenoxyacetic acid (2,4-D) + N-(2-chloro-4-pyridyl)-N'-phenylurea (4-CPPU)). Results were compared with those obtained when explants were cultivated in the presence of 2,4-D + BAP, previously used on melon. Embryogenesis occurred more successfully in 4-day-old cotyledons and seeds than hypocotyls and 8-day-old cotyledons. The best result was achieved with NOA + BAP. Genotypes significantly affected embryogenesis. The number of embryos in "Eivanaki" was significantly higher than that in "Mashhadi." Embryo proliferation when explants were maintained in jars (9.3%) was found to be higher compared to that in petri dishes. For the first time, genetic stability of regenerated melon plants was evaluated using inter-simple sequence repeat markers. Polymerase chain reaction (PCR) products demonstrated a total of 102 well-resolved bands, and regenerants were 93% similar compared to the mother plant. Somaclonal changes during embryogenesis were evaluated by flow cytometry, showing 91% of the same patterns in regenerated plants. The results suggest that the new hormone components are effective when applied for in vitro embryogenesis of muskmelon as they show a high frequency in regeneration and genetic homogeneity.
报道了一种通过直接体细胞胚胎发生实现两个甜瓜品种（甜瓜（Cucumis melo L.）“Mashhadi”和“Eivanaki”）离体再生的新方案。在添加了三种迄今从未在甜瓜上测试过的不同激素组合（萘氧基乙酸（NOA）+噻二唑素（TDZ）、NOA + 6-苄基腺嘌呤（BAP）以及2,4-二氯苯氧乙酸（2,4-D）+ N-(2-氯-4-吡啶基)-N'-苯基脲（4-CPPU））的Murashige和Skoog培养基上培养4日龄和8日龄的子叶、种子及下胚轴，从而获得体细胞胚胎。将结果与在外植体于先前用于甜瓜的2,4-D + BAP存在下培养时所获得的结果进行比较。4日龄子叶和种子中的胚胎发生比下胚轴和8日龄子叶更成功。使用NOA + BAP取得了最佳结果。基因型对胚胎发生有显著影响。“Eivanaki”中的胚胎数量显著高于“Mashhadi”。发现外植体在培养瓶中培养时的胚胎增殖率（9.3%）高于在培养皿中的增殖率。首次使用简单序列重复区间标记评估再生甜瓜植株的遗传稳定性。聚合酶链反应（PCR）产物共显示出102条清晰可辨的条带，再生植株与母本植株的相似度为93%。通过流式细胞术评估胚胎发生过程中的体细胞克隆变异，结果显示再生植株中有91%具有相同模式。结果表明，新的激素成分应用于甜瓜离体胚胎发生时是有效的，因为它们在再生中表现出高频率且遗传同质。
