Monitoring Xylem Transport in Arabidopsis thaliana Seedlings Using Fluorescent Dyes

Fluorescent dyes are often used to observe transport mechanisms in plant vascular tissues. However, it has been technically challenging to apply fluorescent dyes on roots to monitor xylem transport in vivo. Here, we present a fast, noninvasive, and high-throughput protocol to monitor xylem transport in seedlings. Using the fluorescent dyes 5(6)-carboxyfluorescein diacetate (CFDA) and Rhodamine WT, we were able to observe xylem transport on a cellular level in Arabidopsis thaliana roots. We describe how to apply these dyes on primary roots of young seedlings, how to monitor root-to-shoot xylem transport, and how to measure xylem transport velocity in roots. Moreover, we show that our protocol can also be applied to lateral roots and grafted seedlings to assess xylem (re)connection. Altogether, these techniques are useful for investigating xylem functionality in diverse experimental setups.