Band 7.2b is an integral membrane phosphoprotein absent from the erythrocyte membranes of patients with hereditary hydrocytosis, a hemolytic anemia inherited in an autosomal dominant fashion and characterized by stomatocytic red blood cells with abnormal permeability to Na+ and K+. The precise role of band 7.2b is unknown, but it may interact with other proteins of the junctional complex of the membrane skeleton. To gain additional insight into the structure and function of this protein and to provide the necessary tools for further genetic studies of hydrocytosis patients, we determined the sequence of the full-length human band 7.2b cDNA, characterized the genomic structure of the band 7.2b gene, studied its pattern of expression in different tissues, and characterized the promoter of the gene. The composite band 7.2b gene cDNA was 3047 base pairs in length. Northern blot analysis revealed a wide tissue distribution of expression of the band 7.2b gene, with utilization of alternative polyadenylation signals generating transcripts of 2.2 and 3.1 kilobases. Cloning of the band 7.2b chromosomal gene revealed that it is composed of seven exons distributed over 40 kilobases of DNA. The band 7.2b gene promoter was identified as a TATA-less, (G+C)-rich promoter with a typical InR recognition sequence and a single transcription initiation site. It directed high level expression of a reporter gene in both erythroid and nonerythroid cells. An imperfect simple sequence repeat polymorphism was identified in the 5'-flanking DNA, and an assay was developed for its analysis by PCR.