Development and Validation of an SNP-Based OpenArray® Genotyping Panel for Discriminating Coturnix coturnix, Coturnix japonica and Their Hybrids

Background/Objectives: The common quail (Coturnix coturnix) is a game species facing conservation challenges, particularly hybridization with the Japanese quail (Coturnix japonica). To address this issue, one proposed measure is the urgent prohibition of releasing farmed quails into the wild. If authorized, mechanisms should be established to guarantee their genetic origin and prevent contamination of native populations. This work focuses on the development of a genetic tool based on Single-Nucleotide Polymorphism (SNP) markers that can differentiate between the two species and their hybrids. Our goal was to incorporate the selected markers into an OpenArray® platform, to allow efficient, rapid, and cost-effective analysis. Methods: We tested two mitochondrial DNA SNPs (previously described in the literature) as diagnostic markers for species differentiation. We also assessed 24 nuclear DNA SNPs for compatibility with the OpenArray® platform. Results: Of the 26 total SNPs, eight were excluded due to their limited utility. The remaining 18 SNPs achieved an overall genotyping success rate of 96.21%. Using the OpenArray® platform with these 18 SNPs in a trial with samples from diverse Spanish field populations, we found 1.00% of C. japonica alleles (affecting 15.63% of individuals), suggesting introgression in the field. Population genetic analyses revealed strong differentiation between species and confirmed the presence of admixed individuals in field populations. Conclusions: This paper presents a new tool to differentiate between quail species and to identify foreign alleles in stocks and populations, by using an open platform system that optimizes the practical application of the diagnostic procedure based on the to-date most-reliable SNP markers for this goal.