Jul 1, 1976

Naturally occurring cross-links in yeast chromosomal DNA

M A Forte, W L Fangman


Chromosome-size yeast DNA molecules with a number average molecular weight (Mn) of 3-4 X 10(8) were isolated from sucrose gradients after sedimentation of lysed yeast spheroplasts. Resedimentation showed that the molecules were isolated without introducing appreciable single-strand or double-strand breaks. The presence of cross-links in these molecules was suggested by the observation that the apparent Mn in alkali was greater than expected for separated single strands. Since cross-linked molecules would have strands which fail to separate upon denaturation, this was tested more directly. Neutralization of alkaline denaturing conditions resulted in up to 70% of the intact molecules rapidly reforming duplex structures, as shown by equilibrium banding in CsCI. Experiments with larger E. coli DNA molecules (Mn = 5.2 X 10(8)) indicated that the conditions used were sufficient to denature completely molecules of this size. Results of enzyme treatments suggest that the cross-links are not RNA or protein. Experiments with density-labeled yeast DNA molecules showed that the rapid reformation of duplex DNA is not the consequence either of a bimolecular reaction between separated DNA strands or of intrastrand renaturation. The data indic...Continue Reading

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Mentioned in this Paper

Centrifugation, Density Gradient
Nucleic Acid Renaturation
RNA Denaturation
Sedimentation Procedure
Escherichia coli

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