Abstract An outstanding problem in vertebrate development has been to define the genetic program that specifies the cardiomyocyte lineage. It has been a challenge to define the transcription factors that control specification, since candidate gene knockouts typically cause rather complex morphogenetic defects. In contrast, Drosophila genetics identified single transcription factors that are essential for specification of cardiomyocytes from uncommitted mesoderm. For those vertebrate orthologs, it has been considered that paralogous family members might compensate for the loss-of-function of individual genes. However, this hypothesis had not been formally tested. In zebrafish, defects in gata5 can lead to a loss of myocardial tissue, but most embryos depleted for any single vertebrate Gata4/5/6 transcription factor develop a cardiac morphogenetic defect, and cardiomyocytes are specified and differentiate. Here we show that in zebrafish the gata5 and gata6 genes are redundant for specification of cardiomyocytes. Embryos depleted of these two gene products are heartless. Restoring either gene product is sufficient to rescue cardiomyocyte specification. In contrast, embryos depleted of Gata4 and Gata6, or Gata4 and Gata5, develop defective heart tubes. Our study identifies a specific pair of vertebrate transcription factor paralogs that is essential for cardiomyocyte specification.