Polyma DNA: A Physical Map

The action of restriction enzymes on polyoma DNA was studied with uniformly 32 P-labeled viral DNA obtained from either infected 3T6 or secondary mouse-embryo cells. Three restriction enzymes were used to construct a physical map of the polyoma genome. An enzyme from Hemophilus parainfluenzae , Hpa II , cleaved polyma DNA into eight unique fragments (Hpa II -1 to Hpa II -8), ranging in size from 27.3 to 1.8% of the genome. An enzyme from Hemophilus influenzae , Hin III , gave two fragments (56 and 44%); and a third enzyme from Escherichia coli , EcoR I , cut at a single unique site. The physical map of the polyma genome was constructed from methods involving: ( 1 ) further digestion of the fragments produced by enzymes EcoR I and Hin III with Hpa II , and ( 2 ) analysis of the products of partial digestion with Hpa II . Analysis by electron microscopy of replicating DNA molecules (less than 50% replicated) cut with the Hin III enzyme, in combination with other studies, has indicated that the origin of DNA replication is located at 71 ± 3 map units from the EcoR I cleavage site, probably in Hpa II -5.