Tight transcriptional control of foreign genes introduced into the germline of transgenic mice would be of great experimental value in studies of gene function. To develop a system in which the spatial and temporal expression of candidate genes implicated in cardiac development or function could be tightly controlled in vivo, we have generated transgenic mice expressing a tetracycline-controlled transactivator (tTA) under the control of a rat alpha myosin heavy chain promoter (MHC alpha-tTA mice), as well as mice harboring a candidate target gene implicated in the control of differentiation, Id1 (tet-Id1 mice). No expression of the target transgene was detected in any tissues of hemizygous tet-Id1 mice. Genetic crosses with MHC alpha-tTA mice resulted in transactivation of the Id1 transgene, but expression was restricted to heart, where tTA was expressed. Furthermore, transactivation of the target gene was tightly and reversibly controlled by systemic therapy with tetracycline, both in utero and postnatally. These studies demonstrate the feasibility of such a binary approach for tightly controlling the timing and extent of expression of transgenes in vivo. This approach should be generally useful for the ectopic expression of candidate genes in selected tissues during delineated developmental stages.