Shuffling the yeast genome using CRISPR/Cas9-generated DSBs that target the transposable Ty1 elements. Shuffling the yeast genome using CRISPR/Cas9-generated DSBs that target the transposable Ty1 elements

We explored how Cas9-induced double-strand breaks (DSBs) on Ty1 produce genomic alterations in the diploid yeast Saccharomyces cerevisiae. Following Cas9 induction, we observed a significant elevation of chromosome rearrangements (large deletions and duplications), loss of heterozygosity (gene conversions, crossovers, and break-induced replication), and aneuploidy. Almost all of the chromosomal rearrangements reflect the repairing of DSBs at Ty1 elements by homologous recombination. Overall design: The goal of our experiments was to analyze genomic rearrangements induced in yeast by expression of a CRISPR/Cas9 system directed against a target in the repeated transposon Ty1. Using whole genome SNP microarrays, we analyzed genomic alterations in 18 yeast isolates. For each isolate, cells were under long-term or transient (2 hours or 4 hours) exposure of CRISPR/Cas9 for inducing double strand breaks on Ty1. MD741-3, -5, -6, -7 are from long-term exposure. We subcultured MD741-3, -5, -6, -7 for one time to analyze the ongoing instability. 2 colonies from each isolate were picked up for analyzing, named as MD741-3-S1, MD741-3-S2, MD741-5-S1, MD741-5-S2, MD741-6-S1, MD741-6-S2, MD741-7-S1, MD741-7-S2. MD704-2h-7, -26 are from 2 hours exposure. MD704-4h-1, -6P, -8, -12 are from 4 hours exposure.