It has recently emerged that transcriptional differences exist between left and right cardiac chambers. An example is provided by transgenic mice with an nlacZ reporter gene under transcriptional control of the fast skeletal muscle alkali myosin light chain (MLC) 3 promoter and 3' enhancer, which express beta-galactosidase in a left ventricular-right atrial dominant pattern in the developing and adult heart. Here, we demonstrate that endogenous MLC3F transcripts are also left/right regionalised in the mouse heart during embryonic development. Regionalisation is observed as early as embryonic day (E) 8.5, and by E10.5 MLC3F transcripts are present predominantly in the future left ventricle and right atrium, and to a lesser extent in the left atrium. Subsequently, MLC3F transcripts are down-regulated in the left ventricle, and by E12.5 expression is restricted to both atria and left-ventricular trabeculae. No MLC3F protein can be detected in the adult or embryonic mouse heart, suggesting that post-transcriptional regulation prevents this fast myosin isoform contributing to myocardial contraction. Left ventricular-right atrial dominant MLC3F transgenes therefore reflect transitory left/right regionalisation of the endogenous gene, unlike other reported cases of transgene regionalisation. MLC3F transgenes, however, maintain an embryonic-like distribution throughout development suggesting that myocardial gene expression is controlled by distinct temporal, as well as spatial, regulatory modules.