In clinical and pharmacological laboratories, the assay for gamma-glutamyltransferase (GGT) activity is an important diagnostic test, but one with high biological variability. Although the human genome contains multiple GGT genomic sequences, the diagnostic tests generally assume that only a single GGT gene is active. In the current study, segments encompassing parts of seven different potential human GGT genes have been molecularly cloned. Based on sequence determination of exons within these distinct genomic clones, oligonucleotide primers were designed which would prime and PCR-amplify putative mRNA of all seven potential GGT genes, if expressed. Gene-specific oligonucleotide probes were then utilized to assay the transcriptional status of the seven possible GGT genes in a wide variety of human RNAs. Our results show that a single GGT gene exhibits ubiquitous expression in all RNAs tested, including those from fetal and adult liver. A surprisingly large number of four additional GGT genes is expressed in man. Interestingly, these novel GGT genes are expressed in a tissue-restricted manner, which suggests that their corresponding gene products exhibit distinct functions in these specific tissues.