R1-MYB transcription regulatory factors play vital roles in transcriptional modification during higher plant metabolism and development. In the present study, an R1-MYB gene was isolated from the maize (Zea mays L.) inbred line C8605-2 based on expressed sequence tags (ESTs), which expressed differentially between the hybrid C8605-2×W1445 and its two parents in an oligonucleotide microarray. The full-length cDNA, designated as ZmMYBE1 (GeneBank Accession No. FJ024049), consists of 1992 nucleotides and contains a 1296-bp open reading frame. One conserved MYB domain was detected near the N-terminus of the deduced amino acid sequence, where an acidic Ser/Thr-rich area was also observed in the downstream region. The sequence of ZmMYBE1, mapped to chromosome 5 (bin 5.03), was revealed belonging to a multi-gene family in the maize genome. Subcellular localization analysis determined that ZmMYBE1 was located in the nucleus. Expression analysis showed that ZmMYBE1 transcripts accumulated in various tissues examined, with high expression levels in the immature ear and low levels in the tassel, root, and stem. The expression analysis for ZmMYBE1 in immature ears showed the highest expression level at the earliest vegetative developmental stage of the plant. Expression of ZmMYBE1 higher than that of the parents was observed in the hybrid C8605-2×W1445. And also the expression level similar to the lowest of the parents in the hybrid C8605-2×W245 was observed, revealing different expression patterns for the same gene in various hybrids combined from diversified inbred lines. Transgenic Arabidopsis overexpressing ZmMYBE1 implies that ZmMYBE1 has an important role in developmental regulation in maize.