Sites in the polyoma genome cleaved by restriction endonuclease HindII.

Restriction endonucleases are powerful tools with which to study the structure and function of DNA. In particular, endonuclease fragments from SV-40, x and 4x174 DNA’s have provided new insights into viral gene expression and replication (1-7). Recently, the sites of cleavage of a number of restriction endonucleases in polyoma DNA were ordered (8). In this report we add to that list the locations of the two sites cleaved by the Hind11 restriction endonuclease. Pasadena large plaque polyoma 32Plabeled DNA was purified from infected secondary whole mouse embryo cells or mouse 3T6 cells as previously described (9). Care was taken to use low passage virus to minimize the level of defectives (9, 10). Restriction endonucleases were purified from Hemophilus influenzae Rd and H. parainfluenzae by procedures slightly modified from those described by Smith and Wilcox (11) or Sharp, Sugden and Sambrook (12). Digestions were performed at 37” in buffers containing 10 mM Tris-Cl (pH 7.4), 6.6 mM MgCl,, 6 mM mercaptoethanol and 60 mM NaCl (for Hind11 and HindIII) or 10 mM Tris-Cl (pH 7.4), 10 mM MgCl,, 1 mA4 dithiothreitol and 6 mM KC1 (for HpaI and HpaII). Hind11 was separated from Hind111 by chromatograT