Maize has a complex root system composed of different root types formed during different stages of development. The rtcs (rootless concerning crown and seminal roots) mutant is impaired in the initiation of the embryonic seminal roots and the post-embryonic shoot-borne root system. The primary root of the mutant shows a reduced gravitropic response, while its elongation, lateral root density and reaction to exogenously applied auxin is not affected. We report here the map-based cloning of the RTCS gene which encodes a 25.5 kDa LOB domain protein located on chromosome 1S. The RTCS gene has been duplicated during evolution. The RTCS-LIKE (RTCL) gene displays 72% sequence identity on the protein level. Both genes are preferentially expressed in roots. Expression of RTCS in coleoptilar nodes is confined to emerging shoot-borne root primordia. Sequence analyses of the RTCS and RTCL upstream genomic regions identified auxin response elements. Reverse transcriptase-PCR revealed that both genes are auxin induced. Microsynteny analyses between maize and rice genomes revealed co-linearity of 14 genes in the RTCS region. We conclude from our data that RTCS and RTCL are auxin-responsive genes involved in the early events that lead to the initiation and maintenance of seminal and shoot-borne root primordia formation.