A physical map of Anagrapha falcifera multinucleocapsid nuclear polyhedrosis virus (AfMNPV) DNA was constructed for restriction endonucleases EcoRI, HindIII, PstI and XhoI. The genome size was estimated to be 130 kbp. Ordering of the restriction fragments was accomplished by cross-blot hybridization, double digestion and DNA-DNA hybridization. The polyhedrin gene and homologous repeat (hr) regions were located by hybridization to the Autographa californica MNPV (AcMNPV) polyhedrin gene and hr4, respectively. Restriction pattern comparison and Southern blot analysis suggest that AfMNPV is closely related to AcMNPV.