UCSF

At the heart of meiosis is meiotic recombination where programmed double-strand breaks are repaired into either crossovers (COs) or noncrossovers (NCOs). COs promote successful chromosome segregation during the first meiotic division by establishing chiasmata, which are physical connections between homologous chromosomes that provide the tension to properly align chromosomes on the meiosis I spindle. Homologs lacking COs may result in nondisjunction, leading to aneuploid gametes. The number and distribution of COs are tightly regulated to ensure a successful meiotic division. Despite the importance of COs, the mechanisms underlying CO control remain elusive, largely due to the difficulty in determining CO distribution on a genome-wide level.

In this thesis, we describe two methods for mapping the distribution of COs and NCOs genome-wide using two polymorphic Saccharomyces cerevisiae strains, S96 and YJM798. First, we used DNA microarrays to identify ~8000 polymorphic markers in the progeny of S96 and YJM789. Eight meiotic mutants were studied: zip1, zip2, zip3, zip4, msh4, spo16, ndj1, and sgs1. We demonstrated that many aspects of the CO behavior--such as CO level, CO interference, CO homeostasis, chromatid interference, and the behavior of COs near centromeres and telomeres--could be evaluated simultaneously using this method. We showed for the first time that CO homeostasis occurred in wild-type strains. We also identified Zip1 as important for CO suppression at the centromeres.

Using next-generation sequencing, we identified ~54,000 markers and studied the recombination landscape in wild-type and three meiotic mutant tetrads: msh4, sgs1, and pCLB2-MMS4. We demonstrated that next-generation sequencing is a powerful tool for mapping the genome-wide landscape of meiotic recombination events. When coupled with multiplexing, sequencing drastically reduces the cost to lower than that of microarrays, making it possible for large scale experiments involved in studying meiotic mutants. We showed that complex gene conversion motifs near sites of crossing over could be identified and used to unlock the molecular mechanisms and regulations that govern the distribution and formation of recombination events. This technique will prove to be an invaluable contribution to the meiosis field and will help advance our understanding of meiotic recombination in the near future.