Using Xenopus laevis Oocytes to Functionally Characterize Plant Transporters.

Abstract

Functionally characterizing plant membrane transport proteins is challenging. Typically, heterologous systems are used to study them. Immature eggs (oocytes) of the South African clawed frog Xenopus laevis are considered an ideal expression system for such studies. These large oocytes have a low number of endogenous transport systems in their plasma membranes and highly express foreign mRNA; the oocyte plasma membrane is the default destination of integral membrane proteins that lack recognized organellar sorting signals. These features facilitate almost background-free characterization of putative plant membrane transporters. Here we describe how to isolate Xenopus laevis oocytes, prepare capped sense RNA (cRNA) of the maize boron importer TASSEL-LESS1 (TLS1) as an example, microinject the cRNA into the isolated oocytes, and functionally assess the boron import capabilities of TLS1 in an oocyte swelling assay. These protocols can be easily adapted to study other plant and non-plant transporters with putative import function. © 2019 by John Wiley & Sons, Inc.