Dinkeloo Kasia, Cantero Araceli Maria, Paik Inyup, Vulgamott Alexa, Ellington Andrew D, Lloyd Alan
Department of Molecular Biosciences, College of Natural Sciences, The University of Texas at Austin, Austin, TX, 78712, USA.
Plant Methods. 2021 Jun 9;17(1):59. doi: 10.1186/s13007-021-00760-3.
Taraxacum officinale, or the common dandelion, is a widespread perennial species recognized worldwide as a common lawn and garden weed. Common dandelion is also cultivated for use in teas, as edible greens, and for use in traditional medicine. It produces latex and is closely related to the Russian dandelion, T. kok-saghyz, which is being developed as a rubber crop. Additionally, the vast majority of extant common dandelions reproduce asexually through apomictically derived seeds- an important goal for many major crops in modern agriculture. As such, there is increasing interest in the molecular control of important pathways as well as basic molecular biology and reproduction of common dandelion.
Here we present an improved Agrobacterium-based genetic transformation and regeneration protocol, a protocol for generation and transformation of protoplasts using free DNA, and a protocol for leaf Agrobacterium infiltration for transient gene expression. These protocols use easily obtainable leaf explants from soil-grown plants and reagents common to most molecular plant laboratories. We show that common markers used in many plant transformation systems function as expected in common dandelion including fluorescent proteins, GUS, and anthocyanin regulation, as well as resistance to kanamycin, Basta, and hygromycin.
Reproducible, stable and transient transformation methods are presented that will allow for needed molecular structure and function studies of genes and proteins in T. officinale.
药用蒲公英,即普通蒲公英,是一种广泛分布的多年生植物,在全球范围内被视为常见的草坪和花园杂草。普通蒲公英也被种植用于制作茶、作为可食用蔬菜以及用于传统医学。它能产生乳胶,并且与俄罗斯蒲公英(T. kok-saghyz)密切相关,后者正被开发为一种橡胶作物。此外,绝大多数现存的普通蒲公英通过无融合生殖产生的种子进行无性繁殖——这是现代农业中许多主要作物的一个重要目标。因此,人们对普通蒲公英重要途径的分子调控以及基础分子生物学和繁殖的兴趣与日俱增。
在此,我们展示了一种改进的基于农杆菌的遗传转化和再生方案、一种使用游离DNA生成和转化原生质体的方案以及一种用于叶农杆菌浸润以进行瞬时基因表达的方案。这些方案使用从土壤种植的植物中容易获得的叶片外植体以及大多数分子植物实验室常用的试剂。我们表明,许多植物转化系统中使用的常见标记在普通蒲公英中按预期发挥作用,包括荧光蛋白、GUS和花青素调控,以及对卡那霉素、草丁膦和潮霉素的抗性。
本文提出了可重复、稳定和瞬时的转化方法,这将有助于对药用蒲公英中的基因和蛋白质进行所需的分子结构和功能研究。