Location of S1 nuclease-cleavage sites on circular, superhelical DNAs between polyoma virus and simian virus 40.

Physical maps of the S1 nuclease-cleavage sites on double-stranded covalently closed, circular, superhelical DNAs of polyoma virus and simian virus 40 (SV 40) were constructed by the following technique: Polyoma and SV 40 DNAs were digested under defined conditions (30mm sodium acetate buffer (pH 5.0), 60mM NaCl at 37°C) with S1 nuclease in the presence of Mn2+ to generate unit length linear duplex molecules. Then, the linear DNAs were digested with Eco RI and Bam NI restriction endonucleases and size of the resulting DNA fragments were analyzed by agarose gel electrophoresis. Cleavage of those DNAs with S1 is more specific in the presence of Mn2+ than Zn2+ since discrete DNA bands can be obtained after electrophoresis in agarose gels. With Zn2+ a well known activator of the enzyme, S1 cleaves these DNAs within certain regions rather than at specific sites. With Mn2+ as activator S1 cleaves polyoma DNA at seven sites: map coordinate 0.11, 0.21, 0.30, 0.44, 0.56, 0.82 and 0.92 (fractional DNA lengths clockwise from the single Eco RI cleavage site (zero position)). SV 40 DNA is cleaved by S1 under these conditions at map coordinate 0.15, 0.28, 0.38, 0.39, 0.44, 0.56, 0.89 and 0.98 (also defined as fractional DNA lengths clockwise from SV 40's single Eco RI cleavage site (zero position)). Map position 0.21 on polyoma DNA and 0.15 on SV 40 DNA are at or close to the termi-nation site for DNA replication and the 3' ends of early and late mRNAs. Map positions 0.82, 0.92 and 0.11 on polyoma DNA and 0.56, 0.44 and 0.28 on SV 40 DNA occur at about 0.2, 0.4 and 0.8 respectively from the origin of DNA replication in the early region. The DNA segments between map positions 0.44 and 0.56 on polyoma and 0.89 and 0.98 on SV 40 contain the sequence coding for the COOH-terminus of VP2_??_VP3 and the NH4-terminus of VP1.