Isolation and propagation of a segment of the simian virus 40 genome containing the origin of DNA replication

Heteroduplex DNA molecules formed from two DNAs that differ from each other by a deletion can be cleaved at the mismatched region (a deletion loop) with the single strand specific S1 endonuclease. A heteroduplex DNA molecule, constructed from the DNA of a simian virus 40 (SV40) mutant with a deletion of the map region 0.54-0.55 and the DNA of a second SV 40 mutant having a deletion of the map segment 0.70-0.73, is cleaved twice with S1 endonuclease. One of the products is a DNA fragment of about 0.13 the length of SV 40 DNA which contains the origin of SV40 DNA replication (0.67 on the SV40 DNA map). Infection of cultured CV1 monkey kidney cells with the fragment and intact SV40 DNA yields, in addition to the expected full length wild type circular DNA molecules, a population of discrete size circular DNAs whose lengths are very nearly integral multiples of the infecting fragment. Restriction endonuclease digestion patterns and heteroduplex analysis indicate that the small circular DNAs are oligomers of the infecting fragment, organized in 'head to tail' and, less frequently 'head to head' arrangement.