Polyploid QTL-seq identified QTLs controlling potato flesh color and tuber starch phosphorus content in a plexity-dependent manner.

The progenies of polyploid crops inherit multiple sets of homoeologous chromosomes through various combinations, which impedes the identification of the quantitative trait loci (QTL) governing agronomic traits and the implementation of DNA marker-assisted breeding. Previously, we developed a whole-genome sequencing-based polyploid QTL-seq method that utilizes comprehensively extracted simplex polymorphisms for QTL mapping. Here, we verified the detection of duplex QTLs by modifying the analytical settings to explore the QTLs governing tuber flesh color and starch phosphorus content using tetraploid potato (Solanum tuberosum L.). The F₁ progenies were obtained from a cross between 'Touya' (TY) and 'Benimaru' (BM). A single TY-derived QTL responsible for yellow flesh color was identified around a β-carotene hydroxylase gene on chromosome 3 using simplex polymorphisms, and a BM-derived QTL associated with decreased starch phosphorus content near a starch synthase II gene on chromosome 2 was detected using duplex polymorphisms. Furthermore, linked DNA markers were developed at the QTL sites. For the latter QTL, plexity-distinguishable markers were developed using quantitative PCR, fragment analysis, and amplicon sequencing. These revealed the allele dosage-dependent effect of the reduced starch phosphorus content. Thus, the polyploid QTL-seq pipeline can explore versatile QTLs beyond simplex, facilitating DNA marker-assisted breeding in various polyploid crops.