A full-length cDNA encoding the Vicia faba L. early nodulin VfEnod12 was isolated. The deduced protein sequence specified a 90 amino acid protein with a MW of 10206 and contained a putative signal peptide sequence followed by PPX(3) repeats characteristic of Enod12 proteins. The VfEnod12 gene was found to be expressed specifically in root nodules as early as 3 days post inoculation with Rhizobium leguminosarum bv. viciae. In mature nodules, VfEnod12 transcripts were confined to the prefixing zone II. A 3.3 kb genomic fragment carrying the complete VfEnod12 coding region was isolated. No intervening sequences were identified in the coding region. A promoter fragment carrying the -692/-41 region mediated reporter gene expression in root cortical cells, nodule primordia and the prefixing zone II of transgenic Vicia hirsuta root nodules. This fragment contained a putative binding site for the transcription factor ENBP1. In contrast to the highly conserved terminal AATAA motif of the ENBP1 binding site of known Enod12 promoters, the VfEnod12 promoter was characterized by an altered terminal AATAT sequence. This alteration did not interfere with VfEnod12 promoter activity in transgenic roots and nodules of V. hirsuta.