A protocol for mimicking lipid-mediated phase separation on the membrane using giant unilamellar vesicles

In addition to creating a physical boundary, cell membranes provide critical platforms for many biological processes. Due to the complexity of the membrane composition, it is a challenging task to study the functions of membrane-associated proteins. Here, we present a general protocol by which we use giant unilamellar vesicles (GUV) as a tool to study AGO proteins’ behavior on the membrane. We provide procedures to prepare fluorescent-labeled protein and GUVs and describe the protein phase separation in 3D time-lapse images.

除了构建物理边界之外，细胞膜还为众多生物学过程提供了至关重要的平台。鉴于膜组成的复杂性，研究膜相关蛋白的功能是一项颇具挑战性的任务。在此，我们提出一项通用方案，通过利用巨型单层脂质体（GUV）作为研究AGO蛋白在膜上行为的工具。我们提供了制备荧光标记蛋白和GUVs的程序，并在三维时间序列图像中描述了蛋白相分离现象。