Rapid small-scale isolation of SV40 virions and SV40 DNA

A rapid method for the small-scale isolation of SV40 virions and SV40 DNA i s presented. CV-1 monkey epithelial cells are transfected with linear SV40 DNA. After the onset of transfection, cells are lysed by several freeze/tha w cycles and virions are isolated using polyethylene glycol (PEG) precipita tion of DNase I treated lysates. Viral DNA is released by proteinase K and dithiothreitol treatment of the isolated virions followed by phenol/chlorof orm extraction and ethanol precipitation. This method yields on average 7.5 x 10(4) plaque forming units (PFUs) and DNA of adequate purity and concent ration to be used for restriction analysis on ethidium bromide agarose gels from a single 35-mm tissue culture dish. (C) 2000 Elsevier Science B.V. Al l rights reserved.