Development and validation of a DNA test for Silver and Smoke in domestic cats

The Silver and Smoke phenotypes are highly selected coat color traits observed in numerous domestic cat breeds. They are linked to the same Inhibitor (I) locus which creates the Silver effect in Tabby cats and the Smoke effect in Solid (non-Tabby) cats. Phenotypic expression is strongly influenced by underlying coat color genetics and modifying factors, resulting in variation in the extent and distribution of pigment inhibition. Visual assessment can therefore be unreliable in some cats and cannot distinguish heterozygous from homozygous individuals. The Silver locus has previously been linked to a specific interval, but the underlying molecular variant had not been described in the peer-reviewed literature. In this study, comparative screening of WGS data from Silver and non-Silver cats was used to identify putative breakpoints for structural variants associated with the Silver locus. Sequence analysis identified SINE and ERV insertions associated with the Silver locus. A cohort of 83 cats (79 with verified phenotypic data) were genotyped by PCR for both SINE and ERV insertions. ERV genotypes were concordant with all 45 Silver/Smoke and 34 non-Silver cats with high-confidence phenotypes. Phenotypes were associated with the presence of one or both copies of the ERV insertion, which always segregated with the associated SINE element. However, the SINE element was more frequent and was observed in non-Silver cats suggesting a possible pre-existing polymorphism. Genotype results and pedigree analysis support the known dominant mode of inheritance, although larger cohort testing is needed to determine the frequency of alternative Silver-associated variants that may exist. Collectively, these results strongly implicate the ERV insertion as the Silver-associated variant and demonstrate a validated molecular assay for Silver and Smoke genotyping in domestic cats.