Integrating genomics and multi-platform metabolomics enables metabolite QTL detection in breeding-relevant apple germplasm

Research Conducted Apple (Malus × domestica) has commercial and nutritional value, but breeding constraints of tree crops limit varietal improvement. Marker-assisted selection minimizes these drawbacks, but breeders lack applications for targeting fruit phytochemicals. To understand genotype-phytochemical associations in apples, we have developed a high-throughput integration strategy for genomic and multi-platform metabolomics data. Methods 124 apple genotypes, including members of three pedigree-connected breeding families alongside diverse cultivars and wild selections, were genotyped and phenotyped. Metabolite genome-wide association studies (mGWAS) were conducted with 10,000 single nucleotide polymorphisms and phenotypic data acquired via LC-MS and 1H NMR untargeted metabolomics. Putative metabolite quantitative trait loci (mQTL) were then validated via pedigree-based analyses (PBA). Key Results Using our developed method, 519, 726, and 177 putative mQTL were detected in LC-MS positive and negative ionization modes and NMR, respectively. mQTL were indicated on each chromosome, with hotspots on linkage groups 16 and 17. A chlorogenic acid mQTL was discovered on chromosome 17 via mGWAS and validated with a two-step PBA, enabling discovery of novel candidate gene-metabolite relationships. Main Conclusion Complementary data from three metabolomics approaches and dual genomics analyses increased confidence in validity of compound annotation and mQTL detection. Our platform demonstrates the utility of multi-omics integration to advance data-driven, phytochemicalbased plant breeding. ### Competing Interest Statement The authors have declared no competing interest.