Double-strand breaks on artificial chromosomes in yeast

Yeast artificial chromosomes composed primarily of bacteriophage lambda DNA exhibit very low levels of meiotic crossing over compared with similarly s ized intervals of natural yeast DNA. When these recombinationally quiet chr omosomes were augmented with a 12.5 kb insert of sequences from yeast chrom osome VIII, genetic studies demonstrated that the artificial chromosomes ha d acquired recombination properties characteristic of this region of chromo some VIII. On authentic yeast chromosomes, most meiotic recombination event s are initiated at sites where the DNA is cleaved to create a double-strand break (DSB). This report describes physical analyses that were carried out to examine the relationship between DSB sites and the recombination behavi or of the artificial chromosomes. The results show that DSBs are rare on th ese artificial chromosomes, except for the 12.5 kb insert. Mapping of the D SB sites shows that their positions correlate with the previously determine d positions of DSB sites on chromosome VIII. Deletion of two characterized chromosome VIII DSB sites from the 12.5 kb insert on the artificial chromos ome resulted in the loss of the predicted DSB fragments and a reduction in crossing over between artificial chromosomes.