Structure and Replication of SV40 Chromatin
The DNA of SV40 is associated with cellular histones in a chromatin structure in the virions and in the nuclei of infected cells1. Biochemical and electron microscopy studies have shown that viral chromatin, and not pure DNA is the template for the replication machinery in vivo2–7. The process of SV40 replication should thus involve replication of the DNA double strand found in a nucleosomic structure and the doubling of the histone content in the daughter minichromosomes. With the aim of obtaining more information on the structure of the SV40 chromatin, we undertook high resolution electron microscopy studies of viral chromatin8. SV40 minichromosomes were prepared from nuclei of infected cells by the method of Varshavsky et al.9 in the presence of Triton X-100. After purification on sucrose gradients, the chromatin was adsorbed to activated carbon films by the technique of Dubochet et al.10. The samples were briefly stained with aqueous uranyl acetate and observed directly, either by conventional dark field microscopy or with scanning transmission electron microscopy at a magnification of up to x500,000. Figure 1 demonstrates the quality of images obtained by these techniques. Under the conditions employed, the uranyl acetate stains mainly the DNA and not the proteins.
KeywordsInfected Cell Cold Spring Harbor Replicative Intermediate Viral Capsid Protein Aqueous Uranyl Acetate
Unable to display preview. Download preview PDF.
- 1.J. Tooze (ed.), “The molecular biology of tumor viruses”, part 2 DNA tumor viruses, Cold Spring Harbor Laboratory, Cold Spring Harbor, New York, 1980.Google Scholar
- 2.C. Cremisi, P. F. Pignatti, O. Croissant and M. Yaniv. J. Virol. 17, 204, 1976.Google Scholar
- 8.G. Moyne, S. Saragosti and M. Yaniv. J. Mol. Biol. in press.Google Scholar
- 22.S. Saragosti and M. Yaniv, manuscript in preparation.Google Scholar
- 24.P. Herbomel, S. Saragosti, D. Blangy and M. Yaniv. Cell, in press.Google Scholar
- 30.I. Iceckson and M. Yaniv. unpublished observations.Google Scholar
- 38.G. Moyne, F. Harper, S. Saragosti and M. Yaniv. C. R. Acad. Sci. Paris 292, 749, 1981.Google Scholar
- 39.G. Moyne, F. Harper, S. Saragosti and M. Yaniv, in preparation.Google Scholar