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Status |
Public on Apr 29, 2017 |
Title |
Mapping of mitotic gene conversion tracts associated with repair of a defined double-strand break (DSB) using SNP microarray |
Organism |
Saccharomyces cerevisiae |
Experiment type |
Genome variation profiling by SNP array
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Summary |
Mitotic recombination between homologous chromosomes can lead to loss-of-heterozygosity (LOH), which is an important contributor to human disease. In the current study, a defined double-strand break (DSB) on chromosome IV was used to initiate LOH in a yeast strain with sequence-diverged chromosomes. Associated gene conversion tracts, which reflect the repair of mismatches formed when diverged chromosomes exchange single strands, were mapped using microarrays. LOH events reflected two broken chromosomes, one of which was repaired as a crossover and the other as a noncrossover.
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Overall design |
To examine DSB-induced mitotic gene conversion tracts, an I-SceI cut site was inserted into one of the chromosome IVs of a diploid strain (by mating W303 and YJM789 haploid parents) and cleavage by I-SceI enzyme was induced with 45 or 90 minutes of incubation with 2% galactose. Twenty independent DSB-induced red white sectored colonies were analyzed using SNP microarrays specific for yeast chromosome IV. To determine the heterozygous versus homozygous status of SNPs in each event, DNA from individual red or white sectors was labeled with Cy5 and mixed with control parental DNA labeled with Cy3. Following competitive hybridization of the mixed samples to the microarray, the ratio of Cy5 to Cy3 hybridization to each oligo was determined. A hybridization ratio of ~1 to both W303- and YJM879-specific oligos signals heterozygosity for the corresponding SNP. A hybridization ratio >1.5 for a haploid-specific oligo indicates homozygosity for the corresponding SNP, while a ratio <0.5 for the alternative haploid specific indicates loss of the corresponding SNP.
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Contributor(s) |
Hum YF, Jinks-Robertson S |
Citation(s) |
28743762 |
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Submission date |
Apr 28, 2017 |
Last update date |
Jul 25, 2021 |
Contact name |
Yee Fang Hum |
E-mail(s) |
yfhum728@gmail.com
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Phone |
9196845891
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Organization name |
Duke University
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Department |
Molecular Genetics and Microbiology
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Lab |
Jinks-Robertson
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Street address |
381 CARL Building
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City |
Durham |
State/province |
NC |
ZIP/Postal code |
27710 |
Country |
USA |
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Platforms (1) |
GPL21553 |
Agilent-031671 Chr4 RA V2 027289 (SPOT_ID version) |
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Samples (40)
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Relations |
BioProject |
PRJNA384791 |