QTL analysis of high thermotolerance with superior and downgraded parental yeast strains reveals new minor QTLs and converges on novel causative alleles involved in RNA processing

Revealing QTLs with a minor effect in complex traits remains difficult. Initial strategies had limited success because of interference by major QTLs and epistasis. New strategies eliminating major QTLs reduce phenotypic and genetic diversity. Since genetic analysis of superior segregants from natural diploid strains usually also reveals QTLs linked to the inferior parent, we have developed a new approach for minor QTL identification based on elimination of QTLs in both parent strains. We first mapped multiple QTLs by pooled-segregant whole-genome sequence analysis responsible for high thermotolerance in natural yeast strain MUCL28177 compared to BY4742. Using single and bulk reciprocal hemizygosity analysis we identified MKT1 and PRP42 as causative gene in a QTL linked to the superior and inferior parent, respectively. We subsequently downgraded both parents by replacing their superior allele with the inferior allele of the other parent. After crossing, the segregants showed a much wider variation in thermotolerance than the downgraded parent strains, suggesting many more QTLs. Genetic mapping indeed revealed new QTLs and we identified NCS2 and SMD2 as causative genes in two of these. Interestingly, the related functions of PRP42 and SMD2 suggest an important role for RNA processing in thermotolerance. In conclusion, identification of minor QTLs involved in complex traits can be successfully accomplished by crossing parent strains that have both been downgraded for a single QTL. This approach has the advantage of maintaining all relevant genetic diversity and enough phenotypic difference between the parent strains and thus strongly increases the chances of identifying minor QTLs.