A map of temperature-sensitive mutants of simian virus 40.
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Biomedical subjects
Publications and source records attributed to D Nathans.
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Analysis of the viral-specific RNA in simian virus 40(SV40)-infected monkey kidney cells indicated the extensive transcription of both DNA strands. These symmetrically transcribed sequences were localized in the nucleus of infected cells, whereas only the "true" early and late SV40 transcripts were found in the cytoplasm. These results suggest that selective posttranscriptional degradation and/or transport occurs after transcription of the viral DNA. On the basis of hybridization experiments with cytoplasmic RNA and the separated strands of the SV40 Hin fragments, the early SV40 region appears to include all of Hin fragments A, H, I, and B (48% of the genome), whereas the late region is represented in Hin fragments C, D, E, K, F, J, and G (52% of the genome).
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The simian virus 40 (SV40) DNA segments present in a series of adenovirus-SV40 hybrids have been mapped with respect to the sites of cleavage of SV40 DNA by restriction endonucleases. Two approaches have been used. First, nucleic acid hybridizations were performed between equimolar quantities of the denatured DNAs of SV40 and each hybrid virus and the radiolabeled transcripts of 11 DNA fragments obtained by cleavage of SV40 DNA by restriction endonuclease from Hemophilus influenzae. Secondly, selected fragments of SV40 DNA produced by the H. influenzae or H. parainfluenzae restriction endonucleases were used to form heteroduplex DNA molecules with adenovirus and adenovirus-SV40 hybrid DNA, which were then analyzed by electron microscopy. The two sets of data were consistent and have permitted alignment of the map of the SV40 segments of the hybrid viruses with the H. influenzae and H. parainfluenzae cleavage maps of SV40. Since cells infected with some of the hybrid viruses contain one or more SV40-specific antigens, the genetic determinants of these antigens could be localized on the cleavage map.
Serial passage of simian virus 40 (SV40) at high multiplicity of infection leads to the emergence of variants with deleted, substituted, and/or duplicated DNA. Individual variants have been cloned by selective complementation with temperature sensitive SV40 mutants, or nonselectively by coinfection of cells with wild-type helper virus. In each case, the presence of variants was detected by the appearance of discrete short viral genomes in infected cell lysates. Such short genomes, isolated by agarose gel electrophoresis, were shown to be specifically altered by comparing the electrophoretic pattern of their DNA fragments produced by Haemophilus influenzae restriction endonuclease with the pattern of fragments from parental DNA. In addition to defective variants, one infectious variant that had an additional segment of DNA within its genome was isolated.
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