A method for fractionating rat liver cell nuclei on equilibrium density gradients of Cs2SO4
✍ Scribed by Valerio M. Genta; David G. Kaufman; William K. Kaufmann
- Publisher
- Elsevier Science
- Year
- 1975
- Tongue
- English
- Weight
- 661 KB
- Volume
- 67
- Category
- Article
- ISSN
- 0003-2697
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✦ Synopsis
Sonically disrupted nuclei from proliferating liver cells were fractionated in Cs2S0, equilibrium density gradients. Nuclear constituents were concentrated in three bands designated as light band (LB, 1.21 g/cm"), middle band (MB, 1.26 g/cm"), and heavy band (HB, 1.32 g/cm"). Analysis of protein and nucleic acid distribution in gradients suggests preservation of some macromolecular interactions. Studies comparing distributions of radioactively labeled DNA after l-or 120-min intervals following tritiated thymidine injection indicate enrichment of nascent DNA in LB and MB. This enrichment is sensitive to time and pressure of sonication. Furthermore, DNA-polymerase activity was demonstrated in the gradient fractions following removal of Cs,SO1, with most activity once again in the LB and MB. These results suggest this procedure as an initial step in the isolation of an enzymatically active DNA replication complex.