Near-Infrared Autofluorescence Imaging of Nuclei in Living Plant Roots

In live-cell imaging, autofluorescence is often regarded as a negative factor that interferes with the accurate visualization of target fluorescence due to a phenomenon known as crosstalk. However, autofluorescence has also been effectively utilized as an organellar marker. For instance, the intense autofluorescence of chlorophyll in the red wavelength is widely used to visualize chloroplasts, the photosynthetic organelle in plants. Recently, we demonstrated that nuclei in plant cells emit phytochrome-derived autofluorescence in the red to infrared wavelength range, which can be visualized by a conventional confocal microscope equipped with a 640 nm laser. Here, we present protocols for growing plants and conducting confocal imaging of the near-infrared autofluorescence of nuclei in <i>Arabidopsis thaliana</i>. Key features • Less invasive live-cell imaging of plant nuclei using near-infrared (NIR) autofluorescence derived from phytochrome, a photoreceptor in plants. • Fluorescence live-cell imaging of non-model plants. • Imaging of nuclei in root cells and pollen.