Author summary: “The genetic underpinnings of many phenotypic traits in domestic dogs remain undiscovered. Although two genetic loci are known to underlie blue eye color in dogs, these do not explain all cases of blue eyes. By examining > 3,000 dogs…we have shown that a region of canine chromosome 18 carrying a tandem duplication near the ALX4 gene is strongly associated with blue eye color variation, primarily in Siberian Huskies. We also provide evidence that this duplication is associated with blue eye color in non-merle Australian Shepherds.” Read study.