Mitotic sectored colonies: evidence of heteroduplex DNA formation during direct repeat recombination.
In yeast meiosis, ascosporal colonies are sometimes sectored for a marker--i.e., half the colony has one allele and half has the other. This is interpreted as replicative resolution of heteroduplex DNA (hDNA) formed as a recombination intermediate. We have looked for similar evidence of hDNA formation during mitotic recombination between two repeated sequences on the same chromosome. The two repeats, an ochre suppressor and a wild-type tRNA gene, are separated by plasmid DNA and the URA3 marker. Recombination between the repeats excises the URA3 gene and one copy of the repeat, leaving either the wild-type tRNA or the suppressor on the chromosome. A red/white color assay is used to distinguish between the two. We find that some colonies that have lost the URA3 gene are sectored for the suppressor. This suggests that hDNA is formed across the anticodon during the recombination event and then resolved by replication. The disruption of either of two genes involved in recombination and repair, RAD1 and RAD52, does not significantly alter the frequency of sectored colony formation during plasmid excision.
Ronne
H
H
Department of Genetics and Development, College of Physicians and Surgeons, Columbia University, New York, NY 10032.
Rothstein
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R
D002172:Canavanine / Q000378:metabolism; D003470:Culture Media; D004260:DNA Repair; D004271:DNA, Fungal / Q000235:genetics; D005800:Genes, Fungal; D008938:Mitosis; D009963:Orotic Acid / Q000031:analogs & derivatives / Q000378:metabolism; D012343:RNA, Transfer / Q000235:genetics; D011995:Recombination, Genetic; D012091:Repetitive Sequences, Nucleic Acid; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D012641:Selection, Genetic; D013489:Suppression, Genetic
D003470:Culture Media; D004271:DNA, Fungal; D002172:Canavanine; D009963:Orotic Acid; C001242:5-fluoroorotic acid; D012343:RNA, Transfer