Data_Sheet_1_Transformation of Teosinte (Zea mays ssp. parviglumis) via Biolistic Bombardment of Seedling-Derived Callus Tissues.PDF

Modern maize exhibits a significantly different phenotype than its wild progenitor teosinte despite many genetic similarities. Of the many subspecies of Zea mays identified as teosinte, Zea mays ssp. parviglumis is the most closely related to domesticated maize. Understanding teosinte genes and their regulations can provide great insights into the maize domestication process and facilitate breeding for future crop improvement. However, a protocol of genetic transformation, which is essential for gene functional analyses, is not available in teosinte. In this study, we report the establishment of a robust callus induction and regeneration protocol using whorl segments of seedlings germinated from mature seeds of Zea parviglumis. We also report, for the first time, the production of fertile, transgenic teosinte plants using the particle bombardment. Using herbicide resistance genes such as mutant acetolactate synthase (Als) or bialaphos resistance (bar) as selectable markers, we achieved an average transformation frequency of 4.17% (percentage of independent transgenic events in total bombarded explants that produced callus). Expression of visual marker genes of red fluorescent protein tdTomato and β-glucuronidase (gus) could be detected in bombarded callus culture and in T1 and T2 progeny plants. The protocol established in this work provides a major enabling technology for research toward the understanding of this important plant in crop domestication.

尽管现代栽培玉米与其野生祖先大刍草(teosinte)在遗传层面存在诸多相似性，但二者的表型却呈现显著差异。在被归类为大刍草的多个玉蜀黍属(Zea mays)亚种中，小颖大刍草(Zea mays ssp. parviglumis)与栽培玉米的亲缘关系最为紧密。解析大刍草的基因及其调控机制，能够为揭示玉米的驯化历程提供重要见解，并助力未来作物遗传改良育种工作。然而，目前尚无适用于大刍草的遗传转化体系，而该体系是开展基因功能分析的核心技术支撑。本研究以小颖大刍草成熟种子萌发的幼苗叶鞘段为外植体，成功建立了高效稳定的愈伤组织诱导与植株再生体系。本研究同时首次利用基因枪轰击法，获得了可育的转基因大刍草植株。以突变乙酰乳酸合酶(Als)或草丁膦抗性基因(bar)作为筛选标记，本研究实现了平均4.17%的转化效率（即产生愈伤组织的轰击外植体中，独立转基因事件所占的比例）。在轰击后的愈伤组织培养物以及T1、T2代植株中，均可检测到红色荧光蛋白tdTomato与β-葡萄糖苷酸酶(gus)这两类可视化标记基因的表达。本研究建立的转化体系，为解析这一在作物驯化研究中具有重要价值的植物提供了关键的技术支撑。