School of Life Sciences, East China Normal University, Shanghai, China.
Institute of Eco-Chongming, Shanghai, China.
Methods Mol Biol. 2024;2841:225-239. doi: 10.1007/978-1-0716-4059-3_22.
The endomembrane system in plants is composed of interconnected membrane organelles that contribute to intracellular structure and function. These organelles include the endoplasmic reticulum (ER), Golgi apparatus, vacuole, trans-Golgi network, and prevacuolar compartment or multivesicular body. Through vesicle-mediated transport, secreted proteins are synthesized in the ER and subsequently transported along the secretory pathway to the vacuole or outside of cells to fulfill specialized functions. Genetic screening is a crucial method for studying plant protein secretion. It entails identifying phenotypic differences resulting from genetic mutations, such as ethyl methanesulfonate, T-DNA insertion, and RNAi, to investigate gene function and discover mutants with specific traits or gene functions. Significant progress has been achieved in the study of plant protein secretion through genetic screening. In this protocol, we provide a step-by-step guide to studying the protein secretion pathway using a genetic screen approach. We use the example of the free 1 suppressor of Arabidopsis thaliana and oil body mutants of Marchantia polymorpha. Additionally, we offer an overview of genetic screening and briefly summarize the emerging technologies in the field of protein secretion research.
植物的内膜系统由相互连接的膜细胞器组成，这些细胞器有助于细胞内的结构和功能。这些细胞器包括内质网（ER）、高尔基体、液泡、跨高尔基网络和前液泡区室或多泡体。通过囊泡介导的运输，分泌蛋白在内质网中合成，随后沿着分泌途径运输到液泡或细胞外，以完成专门的功能。遗传筛选是研究植物蛋白分泌的重要方法。它涉及识别由于遗传突变（如乙基甲磺酸、T-DNA 插入和 RNAi）引起的表型差异，以研究基因功能并发现具有特定特征或基因功能的突变体。通过遗传筛选，在植物蛋白分泌的研究方面取得了重大进展。在本方案中，我们提供了使用遗传筛选方法研究蛋白分泌途径的分步指南。我们以拟南芥的自由 1 抑制剂和 Marchantia polymorpha 的油体突变体为例。此外，我们概述了遗传筛选，并简要总结了蛋白分泌研究领域新兴技术。
