Agrobacterium tumefaciens is routinely used to engineer desirable genes into dicotyledonous plants. However, the economically important graminaceous plant maize is refractory to tumor induction by inoculation with virulent strains of A. tumefaciens. Currently, the only clearcut evidence for transferred DNA (T-DNA) transport from Agrobacterium to maize comes from agroinfection. To study T-DNA transfer from Agrobacterium to maize cells in a virus-free system, we used here the beta-glucuronidase (GUS; EC 3.2.1.31) gene as a marker. GUS expression was observed with high efficiency on shoots of young maize seedlings after cocultivation with Agrobacterium carrying the GUS gene. Agrobacterium virulence mutants, incapable of transferring T-DNA to dicot tissue, were shown to be deficient in eliciting GUS expression in maize. Hence, expression of the T-DNA-located GUS gene in maize cells is strictly dependent on Agrobacterium-mediated DNA transfer. Histochemical staining of maize shoots revealed GUS expression located mainly in the leaves and the coleoptile.