The location of gene expression of the Agrobacterium tumefaciens ipt gene promoter in transgenic tobacco plants was examined using the β-glucuronidase (GUS) reporter gene. Expression of GUS was detected in every organ and most cell types examined. The highest levels of GUS activity were found in roots. To further examine the transcriptional basis of this broad expression pattern, deletions in the 5′ noncoding region of the gene were translationally fused to two promoterless reporter genes, encoding the enzymes chloramphenicol acetyl transferase (CAT) and β-glucuronidase (GUS). Reporter enzyme assays revealed the existence of an upstream segment required for maximal promoter function, the 5′ end of which is between-442 and-408 of the P_ipt ATG codon. This upstream segment is required for maximal levels of GUS expression in roots, but not in other organs, and a tobacco suspension-cultured cell line. The implications of broad ipt expression on the process of crown gall tumorigenesis are discussed.