Single-molecule imaging has gained momentum to quantify the dynamics of biomolecules in live cells, as it provides direct real-time measurements of various cellular activities under their physiological environment. Yeast, a simple and widely used eukaryote, serves as a good model system to quantify single-molecule dynamics of various cellular processes because of its low genomic and cellular complexities, as well as its facile ability to be genetically manipulated. In the past decade, significant developments have been made regarding the intracellular labeling of biomolecules (proteins, mRNA, fatty acids), the microscopy setups to visualize single-molecules and capture their fast dynamics, and the data analysis pipelines to interpret such dynamics. In this review, we summarize the current state of knowledge for the single-molecule imaging in live yeast cells to provide a ready reference for beginners. We provide a comprehensive table to demonstrate how various labs tailored the imaging regimes and data analysis pipelines to estimate various biophysical parameters for a variety of biological processes. Lastly, we present current challenges and future directions for developing better tools and resources for single-molecule imaging in live yeast cells.

单分子成像已经获得了量化活细胞中生物分子动力学的动力，因为它提供了生理环境下各种细胞活动的直接实时测量。酵母是一种简单且广泛使用的真核生物，由于其低基因组和细胞复杂性以及易于遗传操作的能力，可作为量化各种细胞过程的单分子动力学的良好模型系统。在过去的十年中，在生物分子（蛋白质、mRNA、脂肪酸）的细胞内标记、用于可视化单分子并捕获其快速动态的显微镜设置以及用于解释此类动态的数据分析管道方面取得了重大进展。在这次审查中，我们总结了活酵母细胞单分子成像的当前知识状态，为初学者提供了现成的参考。我们提供了一个综合表格来展示各个实验室如何定制成像方案和数据分析管道，以估计各种生物过程的各种生物物理参数。最后，我们提出了当前的挑战和未来的方向，以开发更好的工具和资源，用于活酵母细胞中的单分子成像。