DNA synthesis in HeLa cell nuclei isolated in a non-aqueous medium
| Autor: | Hans Prydz, Eirik Bjørklid, Unni Spaeren, Carol Sudbery, Knut E. Schrøder |
|---|---|
| Rok vydání: | 1975 |
| Předmět: |
DNA Replication
Glycerol Time Factors Mitosis Cell Fractionation Biochemistry Genetics and Molecular Biology (miscellaneous) Deoxyribonucleotides HeLa chemistry.chemical_compound Centrifugation Density Gradient Humans Thymine Nucleotides Magnesium NADH NADPH Oxidoreductases heterocyclic compounds Centrifugation Cytochrome Reductases Cell Nucleus Differential centrifugation Deoxyribonucleases Chromatography DNA synthesis biology DNA Neoplasm biology.organism_classification Dithiothreitol Microscopy Electron chemistry Biochemistry Ethylmaleimide DNA HeLa Cells Thymidine Homogenization (biology) |
| Zdroj: | Biochimica et Biophysica Acta (BBA) - Nucleic Acids and Protein Synthesis. 395:413-421 |
| ISSN: | 0005-2787 |
| DOI: | 10.1016/0005-2787(75)90065-9 |
| Popis: | Nuclei were isolated from synchronized HeLa cells in the S-phase by a modification of the non-aqueous method described by Kirsch et al. (Science (1970) 168, 1592–1595). The method involved lyophilization of the cells, homogenization in non-aqueous glycerol and centrifugation in a gradient of 0–35% (w/w) 3-chloro-1,2-propanediol in glycerol. Such nuclei incorporated deoxyribonucleotides into DNA when incubated in an aqueous buffer containing Mg2+, ATP, dATP, dGTP, dCTP and dTTP. The product was sensitive to DNAase and banded with bulk DNA in isopycnic centrifugation. Sedimentation of the product in alkaline sucrose gradients after labelling of the nuclei for 2 min revealed labelled material in the 5 S peak and in the 18 S area. The material in the 5 S peak moved into the 12 S area after a 13 min chase. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|