The microbial eukaryote Saccharomyces cerevisiae has 18 chromosomes, each consisting of a DNA molecule of 1 to 15×108 daltons (150 to 2,300 kilobase pairs). Interstand cross-links have now been found in molecules of all sizes by examining the ability of high molecular weight DNA to snap back, i.e., to rapidly renature after denaturation. Experiments in which snap back was assessed for molecules broken by shearing indicate that there are probably two cross-links in each chromosome. Evidence that the cross-links occur at specific sites in the genome was obtained by treating total chromosomal DNA with the endonuclease EcoRI which cleaves the yeast genome into approximately 2,000 discrete fragments. Cross-link containing fragments were separated from fragments without cross-links. This purification resulted in enrichment for about 18 specific fragments. To determine whether the cross-links are terminal or at internal sites in chromosomal DNA, large shear-produced fragments were examined by electron microscopy. With complete denaturation few fragments exhibited the X-shaped single strand configuration expected for internal cross-links. When partially denatured fragments were examined some ends had single strand loops as expected for (AT-rich) cross-linked termini. The percentage of looped ends was sufficient to account for all the cross-links in the population of chromosomal molecules. The data suggest that yeast chromosomal DNA molecules have cross-linked termini. We propose that a duplex chromosomal DNA molecule in this eukaryote consists of a continuous, single, self-complementary strand of DNA. This structure has implications for the mechanism of chromosome replication and may be the basis of telomere behavior.
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