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. 1986 Dec 9;14(23):9457–9470. doi: 10.1093/nar/14.23.9457

Simian virus 40 DNA replication in vitro: purification and characterization of replication factors from mouse cells.

H Ariga
PMCID: PMC311970  PMID: 3025815

Abstract

We have previously developed simian virus 40 (SV40) DNA replication system in vitro (Ariga and Sugano, J. Virol. 48, 481, 1983). This system is composed of human HeLa or mouse FM3A nuclear extract and cytoplasmic extract of SV40 infected CosI cells. Here FM3A nuclear extract was fractionated by DEAE Sephacel and single-stranded DNA cellulose chromatography into three components required for accurate in vitro SV40 DNA replication. One fraction (A fraction) contained DNA polymerase-primase, and the second component (B fraction) contained DNA topoisomerase. Third component was further purified to near homogenuity using DEAE-Sephacel, single-stranded DNA cellulose, and glycerol gradient centrifugation. The purified protein (named factor I) bound to the origin containing fragment of SV40 DNA. The factor I enhanced the initiation of SV40 DNA replication catalyzed by SV40 infected CosI cytoplasm alone. When all four fractions consisting of A, B fractions, factor I, and SV40 infected CosI cytoplasm were mixed together, the system was reconstituted, meaning that initiation and subsequent elongation were completed to generate the full sized daughter molecules.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

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