Even though stable genomic transformation of sporelings and thalli of Marchantia polymorpha is straightforward and efficient, numerous problems can arise during critical phases of the process such as efficient spore production, poor selection capacity of antibiotics or low transformation efficiency. It is therefore also desirable to establish quick methods not relying on stable transgenics to analyze the localization, interactions and functions of proteins of interest. The introduction of foreign DNA into living cells via biolistic mechanisms has been first reported roughly 30 years ago and has been commonly exploited in established plant model species such as Arabidopsis thaliana or Nicotiana benthamiana. Here, we report the fast and reliable transient biolistic transformation of Marchantia thallus epidermal cells using fluorescent protein fusions. We present a catalog of fluorescent markers which can be readily used for tagging of a variety of subcellular compartments. Moreover, we report the functionality of the bimolecular fluorescence complementation (BiFC) in M. polymorpha with the example of the p-body markers MpDCP1/2. Finally, we provide standard staining procedures for live cell imaging in M. polymorpha, applicable to visualize cell boundaries or cellular structures, to complement or support protein localizations and to understand how results gained by transient transformations can be embedded in cell architecture and dynamics. Taken together, we offer a set of easy and quick tools for experiments that aim at understanding subcellular localization, protein–protein interactions and thus functions of proteins of interest in the emerging early diverging land plant model M. polymorpha.

尽管孢子体和菌体的基因组稳定转化地钱虽然简单且高效，但在该过程的关键阶段可能会出现许多问题，例如有效的孢子产生、抗生素的选择能力差或转化效率低。因此，还需要建立不依赖于稳定转基因的快速方法来分析感兴趣蛋白质的定位、相互作用和功能。大约 30 年前首次报道了通过生物射弹机制将外源 DNA 引入活细胞，并且已在已建立的植物模型物种中得到普遍利用，例如拟南芥或者本塞姆氏烟草。在这里，我们报告了使用荧光蛋白融合对地钱菌体表皮细胞进行快速、可靠的瞬时基因枪转化。我们提供了一系列荧光标记物，可轻松用于标记各种亚细胞区室。此外，我们报告了双分子荧光互补（BiFC）在地钱以 p 体标记 MpDCP1/2 为例。最后，我们提供活细胞成像的标准染色程序地钱，适用于可视化细胞边界或细胞结构，补充或支持蛋白质定位，并了解如何将瞬时转化获得的结果嵌入到细胞结构和动力学中。总而言之，我们提供了一套简单快捷的实验工具，旨在了解亚细胞定位、蛋白质-蛋白质相互作用，从而了解新兴早期分化陆地植物模型中感兴趣的蛋白质的功能地钱。