Simultaneous identification of CSN3 and LGB genotypes in cattle by high-resolution melting curve analysis

Genetic polymorphisms of bovine κ-casein (CSN3) and β-lactoglobulin (LGB) have been shown to affect milk compositional traits and technological properties. Monitoring individual CSN3 and LGB genetic variants in the population is important in the subsequent process of breeding cattle and improving milk protein quality. The objective of this study was to develop and validate a method for genotyping the main mutations of the bovine CSN3 and LGB genes using real time PCR (RT-PCR) followed by a high-resolution melting method (HRM) assay. The HRM-based assay allowed the sensitive and fast identification of the CSN3 AA, AB, BB, AE and BE genotypes and LGB AA, AB and BB genotypes simultaneously. This method was validated by analyzing 100 Fleckvieh cattle DNA samples, which were also genotyped by conventional methods at the molecular level using PCR-RFLP and DNA sequencing techniques. The genotypes obtained using the HRM approach perfectly matched the genotypes obtained by conventional analyses. Moreover, it is a cheap and fast alternative for existing methods.