Abstract
BACKGROUND: Over the past thirty years, the identification of species-specific molecular markers has significantly advanced our understanding of genetic diversity in both plants and animals. Among these, short InDels have emerged as vital genomic features, contributing more to sequence divergence than single nucleotide polymorphisms do in closely related species. This study aimed to identify specific InDels for Bos taurus, Bubalus bubalis, Capra hircus, and Ovis aries via an in silico approach and validated them in 400 individuals (100 for each species). RESULTS: We identified and characterized short, specific InDels in the sequences of the CSN1S1, CSN1S2, MSTN, and PRLR genes, which can be used for species identification of Capra hircus, Ovis aries, Bos taurus, and Bubalus bubalis, respectively. We developed a Tetraplex Specific PCR assay to enable efficient discrimination among these species. CONCLUSIONS: This study highlights the utility of InDels as biallelic, codominant markers that are cost-effective and easy to analyse, providing valuable tools for genetic diversity analysis and species identification.