Brachyury (T) is involved in mesoderm induction during early mouse development. T expression in embryonal carcinoma P19 cells, which differentiate into mesoderm derivatives in vitro, was studied. Endogenous T expression in P19 cells was transiently induced when the cells were allowed to form aggregates. This expression was enhanced by dimethyl sulfoxide. In situ hybridization showed that T expressing cells formed clusters on the aggregates. Transfection of plasmids encoding reporter genes under the control of the upstream region of T showed that the sequence up to -351 bp can resemble the differentiation-dependent expression of T in P19 cells. To define the promoter region regulating T expression, transgenic mice carrying LacZ under the control of the upstream region were prepared. The region up to -351 bp is sufficient to direct the expression in the primitive streak and tail-bud. The upstream region up to -2400 bp does not support expression in the notochord. The sequence between -987 and -585 bp enhances expression in the primitive streak and tail-bud. In the tail-bud where new cells for elongation of the anteroposterior axis were continuously supplied, the sequence up to -987 bp drove lacZ expression in gut endoderm and prospective neuroectoderm as well as in mesoderm derivatives.