Novel LNA probe-based assay for the A1 and A2 identification of β-casein gene in milk samples.

The rising consumption of milk and its derivatives in recent years has garnered attention from both consumers and producers, mainly due its possible health benefits, such as enhanced digestion and easier absorption. Thus, a novel real-time PCR using a combination of locked nucleic acid modified (LNA) conjugated probes was developed to genotype and alleles of -casein gene () and to detect and quantify the presence in samples. The limit of detection for each probe ( and ) was evaluated using decreasing serial dilutions. Besides, the sensitivity of A1 allele detection in the samples was also tested. The limits of detection of and alleles were 6 copies, while for allele detection in samples was 7.5 copies (1%). The LNA-probe based method was found to be rapid, robust, highly sensitive, cost effective, and can be employed as screening test to certificate the dairy products.