Gammaherpesvirus (${gamma}HV$) infection of the central nervous system (CNS) has been implicated in diverse neurological diseases, and murine ${gamma}HV$-68 (MHV-68) is known to persist in the brain after cerebral infection. The underlying molecular mechanisms of persistency of virus in the brain are poorly understood. Here, we characterized a unique pattern of MHV-68 persistent infection in neuroblastoma cells. On infection with MHV-68, both murine and human neuroblastoma cells expressed viral lytic proteins and produced virions. However, the infected cells survived productive infection and could be cultured for multiple passages without affecting their cellular growth. Latent infection as well as productive replication was established in these prolonged cultures, and lytic replication was further increased by treatment with lytic inducers. Our results provide a novel system to study persistent infection of ${gamma}HVs$ in vitro following de novo infection and suggest application of MHV-68 as a potential gene transfer vector to the brain.