The fluorescent probe DISBAC2(3) provides a high-throughput screening tool for evaluating abiotic stress tolerance in plants.

The electric gradient across cellular membranes (termed membrane potential [MP]) provides a driving force for the uptake and translocation of all essential nutrients in their ionic form, as well as operating in stress sensing. MP is causally associated with abiotic stress tolerance in plants and, thus, could be used as a proxy in phenotyping programs. However, the conventional method to detect MP changes, which involves impaling the membrane with a microelectrode, requires specialized equipment and specialist skills, is highly time consuming, and is prone to various possible artifacts. Here, we present a high-throughput screening approach that enables the rapid detection of MP changes in plants using the fluorescent probe DISBAC2(3). Using 3 case studies (salinity stress and hypoxia stress in roots and light fluctuations in guard cell movements), we benchmarked this method against conventional microelectrode impalements and demonstrated the feasibility of using this dye for a high-throughput MP screening in various cell types of different species in response to different abiotic stimuli. Through these studies, we show that a voltage-sensitive probe can rapidly and effectively measure the changes in MP elicited by fluctuating environments. We emphasize that the use of such techniques in breeding programs (rather than the time-consuming conventional methods) provides a solution to rapidly identify stress-resistant crops.