Abstract Clinical mastitis (CM) is an inflammatory disease occurring in the mammary glands of lactating cows. CM is under genetic control, and a prominent CM resistance QTL located on chromosome 6 was reported in various dairy cattle breeds. Nevertheless, the biological mechanism underpinning this QTL has been lacking. Herein, we mapped, fine-mapped, and discovered the putative causal variant underlying this CM resistance QTL in the Dutch dairy cattle population. We identified a ~ 12 kb multi-allelic copy number variant (CNV), that is in perfect linkage disequilibrium with a GWAS lead SNP, as a promising candidate variant. By implementing a genome-wide association study (GWAS) and through expression QTL mapping, we showed that the group-specific component gene ( GC ), a gene encoding a vitamin D binding protein, is an excellent candidate causal gene for the QTL. The multiplicated alleles are associated with increased GC expression and low CM resistance. Ample evidence from functional genomics data supports the presence of an enhancer within this CNV, which would exert cis -regulatory effect on GC . We observed that strong positive selection swept the region near the CNV, and haplotypes associated with the multiplicated allele were strongly selected for. Moreover, the multiplicated allele showed pleiotropic effects for increased milk yield and reduced fertility, hinting that a shared underlying biology for these effects may revolve around the vitamin D pathway. These findings together suggest a putative causal variant of a CM resistance QTL, where a cis -regulatory element located within a CNV can alter gene expression and affect multiple economically important traits. Author summary Clinical mastitis (CM) is an inflammatory disease that negatively influences dairy production and compromises animal welfare. Although one major genetic locus for CM resistance was mapped on bovine chromosome 6, a mechanistic description of this association has been lacking. Herein, we report a 12-kb multiallelic copy number variant (CNV), encompassing a strong enhancer for group-specific component gene ( GC ), as a likely causal variant for this locus. This CNV is associated with high GC expression and low CM resistance. We speculate that upregulation of GC leads to a large amount of vitamin D binding protein, which in turn, reduces biologically available vitamin D, resulting in vitamin D deficiency and low CM resistance. Despite the negative effect on CM resistance, the CNV contributes to increased milk production, hinting at balancing selection. Our results highlight how multiplication of a regulatory element can shape economically important traits in dairy cattle, both in favourable and unfavourable directions.

A 12 kb multi-allelic copy number variation encompassing a GC gene enhancer is associated with mastitis resistance in dairy cattle