Triggering of Ag receptors on lymphocytes induces rapid phosphorylation of several receptor-associated protein tyrosine kinases (PTKs), implicating their role in controlling cellular growth and differentiation. In this study, we report the cloning of a human cDNA encoding a nonreceptor PTK with a calculated M(r) of about 58 kDa. The kinase has an overall amino acid identity of approximately 87% with the murine Blk. However, in the unique domain there is only 58% homology and an insertion of six amino acids in the N-terminal region. The nature of this insertion suggests a functional role in membrane attachment. Northern blot analysis showed expression in all stages of B cell development and in T cell lines. The message was not observed in the nonlymphoid tissues examined. In contrast, expression of murine blk in plasma cells and T lymphocytes has not been reported. Importantly, transcripts were seen in human embryonic liver as early as 7.5 wk of gestation before the rearrangement of Ig H chain locus. Furthermore, transcripts were detected in human thymocytes and not in mature T cells. Southern blot analysis revealed polymorphism of this gene in a Caucasian population but not in a Gambian population, indicating a recent origin of this polymorphism. The gene was localized to chromosome 8p22-23. The homology at the protein level suggests that this kinase may be the human homologue of murine Blk. Expression of BLK in immature T cells suggests that BLK may play an important role in thymopoiesis.