Anticarsia gemmatalis multicapsid nucleopolyhedrovirus (AgMNPV) is one of the most successful biological insecticides. In this study, we cloned and sequenced a 12.5 kbp BamHI-D restriction endonuclease fragment of the AgMNPV isolate 2D genome that includes the gp64 gene. We compared this highly conserved region with that of other baculoviruses. AgMNPV contained two genes, p22.2 and v-trex, in common with Choristoneura fumiferana MNPV (CfMNPV) that were not present in other baculoviruses. The v-trex gene has homology to a eukaryotic 3' repair exonuclease and appears to have been acquired from an invertebrate host. The v-trex gene product has the potential to be involved in virus recombination or UV-light tolerance. Multigene phylogenetic analysis suggested that AgMNPV is most closely related to Orgyia pseudotsugata MNPV (OpMNPV). AgMNPV differed from other group I NPVs in that ChiA and v-cath gene homologues were missing from the region downstream of the gp64 gene. Proteinase assays and genetic probes suggest the v-cath gene is absent from AgMNPV.