Beta-casein (CSN2) constitutes up to 45% of the casein of bovine milk. Among the 12 known genetic variants of β-casein gene, the most common forms of β-casein in dairy cattle breeds are A1 and A2 β-casein variants. The present study aims to study the β-casein genotypes of the milk samples (506) that were received at TRPVB facility from various organized farms of Tamil Nadu. This study included native cattle breed (139) and cross bred (367) cattle. The DNA was extracted from samples received on whatman cards and the genotype was determined using allele specific PCR. The results of the study showed that the genotypic frequency for A1/A1, A2/A2 and A1/A2 of native cattle breed was 0%, 78% and 22%, respectively. While the genotypic frequency for A1/A1, A2/A2 and A1/A2 of cross bred cattle was 4%, 63% and 33%, respectively. The gene frequency of β-casein in crossbred cattle was 20% (A1) and 80% (A2); whereas the native cattle breed exhibited A1 allele frequency of 11% and A2 allele frequency of 89%.