Human cytomegalovirus (HCMV) is a human pathogen that causes severe disease primarily in the immunocompromised or immunologically immature individual. To date, no vaccine is available. We describe use of a spread-deficient murine CMV (MCMV) as a novel approach for betaherpesvirus vaccination. To generate a spread-deficient MCMV, the conserved, essential gene M94 was deleted. Immunization with MCMV-ΔM94 is apathogenic and protective against wild-type challenge even in highly susceptible IFNαβR^−/− mice. MCMV-ΔM94 was able to induce a robust CD4⁺ and CD8⁺ T-cell response as well as a neutralizing antibody response comparable to that induced by wild-type infection. Endothelial cells were identified as activators of CD8⁺ T cells in vivo. Thus, the vaccination with a spread-deficient betaherpesvirus is a safe and protective strategy and allows the linkage between cell tropism and immunogenicity. Furthermore, genomes of MCMV-ΔM94 were present in lungs 12 months after infection, revealing first-target cells as sites of genome maintenance.