Gene mapping methodology powered by induced genome rearrangements

Phenotypic variation occurs through genome rearrangements and mutations in certain responsible genes; however, systematic gene identification methodologies based on genome rearrangements have not been fully established. Here, we explored the loci responsible for the given phenotype using the TAQing system and compared it with a conventional mutagenesis-based method. Two yeast strains with different genetic backgrounds and flocculation phenotypes were fused and genomic rearrangements were induced by transient DNA breaks. Then, selection pressure was applied and multiple mutants were generated, showing different flocculation abilities. We also raised mutants with altered cohesiveness due to spontaneous mutations during long-term recursive passages of haploid strains without TAQing treatment. Comparative genomic analysis of the TAQed mutants revealed three chromosomal regions harboring pivotal flocculation genes, whereas conventional mutagenesis generated a more diverse list of candidate loci after prolonged selection. The combined use of these approaches will accelerate the identification of genes involved in complex phenotypes.