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A defined molecular-weight distribution of deoxyribonucleic acid after extensive sonication.

ABSTRACT: Concentrated solutions of low-molecular-weight DNA (Mw=35000) with a known molecular-weight distribution can be prepared in several hours, and require no additional fractionation procedures. This is achieved by sonication of the DNA in 1.0 M-NaCl at high power at 0--2 degrees C. No denaturation of the DNA is detectable, even after 8h of continuous sonication. After 2h, the molecular-weight distribution of the total DNA sample is that of the most probably Schulz distribution, described by-Mn:-Mw:-Mz ...=1:2:3 ...etc. Such a molecular-weight distribution is expected for a random break-up of indefinitely long macromolecules and indicates that the sonication process is essentially by random double-strand scission. DNA was also sonicated in the presence of ligands capable of modifying the DNA tertiary structure. The results support the idea that inflexibility of the DNA is required for efficient sonic degradation.

SUBMITTER: Davis AW

PROVIDER: S-EPMC1185760 | biostudies-other | 1978 Jul

REPOSITORIES: biostudies-other

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Deoxyribonucleic acid synthesis in mammalian systems. Permealysed Ehrlich ascites cells in vitro first label Okazaki-type low-molecular-weight deoxyribonucleic acid and then high-molecular-weight deoxyribonucleic acid.

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Project description:Nuclear and cytoplasmic fractions were prepared from exponentially-growing BHK-21/C13 cells; DNA polymerase was extracted from them and analysed by gel filtration and sucrose-density-gradient centrifugation. DNA polymerase I is heterogeneous comprising species covering a considerable range of molecular weights. These have been tentatively identified as four subspecies of apparent molecular weights 900000-1000000 (IA), 460000-560000 (IB), 270000-320000 (IC) and 140000-200000 (ID), as assessed by gel filtration through Sepharose 6B. DNA polymerase II has a mol.wt. of 46000 +/- 4000 as assessed by gel filtration on Sepharose 6B, and 48000 +/- 2000 as assessed by gel filtration on Sephadex G-100. Sedimentation analyses on sucrose density gradients showed that the DNA polymerase I species had sedimentation coefficients predominantly in the range 6-8 S. DNA polymerase II had predominantly a sedimentation coefficient of 3.2 S although a component with lower sedimentation coefficient was found. The lack of correlation between the molecular weights derived from gel filtration and the sedimentation coefficients is attributed to molecular asymmetry. DNA polymerase I was found to be associated predominantly with the cytoplasm although certain types of nuclear preparation contained large amounts of it. DNA polymerase II was found to be mostly if not exclusively in nuclear preparations.

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