In an effort to misexpress genes in zebrafish, we tested the ability of baculovirus to infect and drive gene expression in embryos. By injecting virus into specific tissues and using appropriate promoters, both the location and time of gene expression could be controlled. Using a virus with 2 different promoters, LacZ and GFP could be expressed independently. The efficiency of expression appears to depend on the promoter used. As a test of this system, baculovirus was used to ectopically express ephrinB2a in the presomitic mesoderm. EphrinB2a is normally expressed in the posterior region of developing somites, and baculovirus-mediated misexpression caused abnormal somite boundary formation. Baculovirus can thus be used as a tool for gene misexpression experiments in the zebrafish, especially when localized misexpression is required late in development.