Purayil Fayas Thayale, Alzaabi Mariam, Sasi Shina, Krishnan Saranya, Badar Zarreen, Li Ling, Kottackal Martin, Amiri Khaled M A
Khalifa Center for Genetic Engineering and Biotechnology, Al Ain, United Arab Emirates.
Research Institute for Science and Engineering, University of Sharjah, Sharjah, United Arab Emirates.
Physiol Plant. 2025 May-Jun;177(3):e70257. doi: 10.1111/ppl.70257.
Improvement of leafy greens, especially perennials with year-round harvesting, is binding to the food security drive. "Food for All" by WHO demands the improvement of regional crops due to the agroclimatic specificity to ensure regional food security. Water spinach (Ipomoea aquatica) is a perennial nutritious leafy green with regional/ethnic cultivation. We accomplished organogenesis and somatic embryogenesis from different explants of I. aquatica, and transgenesis and genome editing through Agrobacterium-mediated transformation. The Ipomoea Basal (CLC-CP) medium was superior to the Murashige and Skoog medium. Hypocotyl explants produced a mean of 12.4 shoots on CLC-CP containing 4.5 μM thidiazuron and 8.7 μM gibberellic acid (GA), 50 mg l ascorbic acid (AA), and 100 mg l adenine hemi-sulfate (AdS). Leaf and root explants induced the highest somatic embryos on a medium containing AdS, AA, 4.4/4.7 μM 6-benzyladenine/kinetin (KIN), and 0.45 μM 2,4-dichlorophenoxyacetic acid. CLC-CP medium with 4.7 μM KIN, 8.7 μM GA, AA, and AdS exhibited elongation of hypocotyl-derived shoots and maturation of somatic embryos. A. tumefaciens-mediated transformation of hypocotyl developed a mean of 3.7 GFP expressing shoots per explant; leaf and root produced 4.3 and 3.1 somatic embryos, respectively. A. rhizogenes infection induced a mean of 4.1 and 3.4 hairy roots from leaf and root explants, respectively. Western blotting of the GFP protein validates water spinach to express human therapeutic proteins. Genome editing of IaNAP1 using hypocotyl explants confirmed the reproducibility of transformation. The plantlets exhibited 100% survival in soil. The present protocol is useful for improving this ethnic leafy green with traits-of-interest.
改善叶菜类蔬菜，特别是可全年收获的多年生叶菜类蔬菜，对粮食安全运动至关重要。世界卫生组织的“人人享有食物”倡议要求根据农业气候特性改良区域作物，以确保区域粮食安全。蕹菜（Ipomoea aquatica）是一种多年生营养叶菜类蔬菜，有区域/民族种植。我们通过蕹菜的不同外植体实现了器官发生和体细胞胚胎发生，并通过农杆菌介导的转化进行了转基因和基因组编辑。蕹菜基础（CLC-CP）培养基优于Murashige和Skoog培养基。在含有4.5 μM噻苯隆、8.7 μM赤霉素（GA）、50 mg l抗坏血酸（AA）和100 mg l腺嘌呤半硫酸盐（AdS）的CLC-CP培养基上，下胚轴外植体平均产生12.4个芽。在含有AdS、AA、4.4/4.7 μM 6-苄基腺嘌呤/激动素（KIN）和0.45 μM 2,4-二氯苯氧乙酸的培养基上，叶片和根外植体诱导出最高数量的体细胞胚胎。含有4.7 μM KIN、8.7 μM GA、AA和AdS的CLC-CP培养基使下胚轴来源的芽伸长并使体细胞胚胎成熟。农杆菌介导的下胚轴转化每个外植体平均产生3.7个表达绿色荧光蛋白（GFP）的芽；叶片和根分别产生4.3个和3.1个体细胞胚胎。发根农杆菌感染分别从叶片和根外植体诱导出平均4.1条和3.4条毛状根。GFP蛋白的蛋白质免疫印迹法验证了蕹菜可表达人类治疗性蛋白质。使用下胚轴外植体对IaNAP1进行基因组编辑证实了转化的可重复性。这些小植株在土壤中的存活率为100%。本方案对于改良这种具有感兴趣性状的民族叶菜类蔬菜很有用。
