Method for fast staining and obtaining high-magnification and high-resolution cell images of Nicotiana benthamiana

As tools of plant molecular biology, fluorescence microscopy and Nicotiana benthamiana have been used frequently to study the structure and function of plant cells. However, it is difficult to obtain ideal micrographs; for example, the images are typically unclear, the inner cell structure cannot be observed under a high-power lens by fluorescence microscopy, etc. Here, we describe a method for observing the cell structure of N. benthamiana. This method significantly improves imaging by fluorescence microscopy and allows clear images to be obtained under a high-power lens. This method is easy to perform with good stability, and the stomatal structure, nucleus, nucleolus, chloroplast and other organelles in N. benthamiana cells as well as protein localizations and the locations of protein–protein interactions have been observed clearly. Furthermore, compared with traditional methods, fluorescent dye more efficiently dyes cells with this method. The applicability of this method was verified by performing confocal scanning laser microscopy (CSLM), and CSLM imaging was greatly improved. Thus, our results provided a method to visualize the subcellular structures of live cells in the leaves of N. benthamiana by greatly improving imaging under a fluorescence microscope and provided new insights and references for the study of cell structures and functions in other plants.