User Manual for the Fluorescent Biosensor nlsNiMet3.0 for Nitrate Ion Detection

In 2022, Dr. Cheng-Hsun Ho and his laboratory team published the first fluorescent resonance energy transfer (FRET) sensor for detecting nitrate ions (NO3–) within plant organisms in the journal Science Advances (Full article link: https://www.science.org/doi/10.1126/sciadv.abq4915). To facilitate a better understanding of how to use the sensor and its potential applications, an experimental protocol and applications for the NiMet3.0 sensor were provided in August 2023 through Bio-protocol.

Nitrate (NO3–) is an essential element and nutrient for plants and animals. However, our knowledge of the distribution of different nitrogen forms in various root cell types and their cellular components remains limited. To address this issue, we have developed a fluorescent biosensor that is targeted to the cell nucleus and composed of genetically encoded components. This sensor is not only reversible but also highly specific, and we have named it nlsNitraMeter3.0. In this manual, we provide a detailed description of how to visualize and quantify nitrate ion concentrations and distributions in Arabidopsis at the cellular level using fluorescence imaging systems such as confocal microscopy.

In addition to guiding readers through the steps from DNA construction to quantification, calculation, and interpretation of imaging results, we also provide supplementary information on important considerations and possible extensions. The tools developed by our team can be of significant assistance in plant biology research, particularly in the study of nitrogen elements.

(For the complete content, please refer to the following link: https://bio-protocol.org/en/bpdetail?id=4743&type=0)

Cheng-Hsun Ho

Yen-Ning Chen and Cheng-Hsun Ho* (2023) User Manual for the Fluorescent Biosensor nlsNiMet3.0 for Nitrate Ion Detection Bio-protocol, 13(16): e4743. DOI: 10.21769/BioProtoc.4743