Cloning and characterization of the integrated viral DNA from three lines of SV40-transformed mouse cells

Abstract

Recombinant DNA techniques have been used to isolate the integrated viral DNA, together with flanking cellular sequences, from three independently isolated lines of SV40-transformed BALB/c 3T3 cells. We have determined the structures of these cloned segments and their relationships to one another by restriction enzyme mapping, electron microscopic heteroduplex analysis and transfer hybridization experiments. Our data indicate that the structure of the integrated viral DNA is consistent with the idea that integration involves a tandemly repeated intermediate, that extensive amplification and rearrangement of viral and adjacent cellular sequences occurs after integration and that the majority of viral DNA insertions do not contain an intact early transcription unit. The altered templates presumably encode the truncated and super T antigens found in these cells. Either they are interrupted in the middle of the early region, in which case they generally retain the region in which tsA mutations map, or they contain tandem duplications of the tsA region. Further structural analysis will allow the prediction of the precise structures of these unusual T antigens, and the availability of the corresponding templates opens the way to an analysis of the functions of these proteins.