Kappa casein plays a crucial role in the formation of stable casein micelles and has a key influence on milk-clotting properties. However, current understanding of buffalo 3 gene polymorphisms is not sufficient. In this study, the polymorphisms in the complete coding sequence (CDS) of the buffalo 3 were detected using PCR product direct sequencing. The CDS of for river and swamp buffalo was the same in length, which contained an open reading frame of 573 nucleotides encoding a peptide containing 190 amino acid residues. A total of eight single nucleotide polymorphisms (SNPs) was identified in two types of buffalo. Among them, c.86C>T, c.252G>C, c.445G>A, c.467C>T and c.516A>C were non-synonymous, which leads to p.Pro8Leu, p.Lys63Asn, p.Val128Ile, p.Thr135Ile and p.Glu151Asp substitutions in buffalo kappa casein ( -CN), respectively. The substitution of p.Thr135Ile may exert a vital effect on the function of buffalo -CN. Eleven haplotypes were defined based on the SNPs found in buffalo, and accordingly, seven protein variants and four synonymous variants of buffalo -CN were inferred, called variants A, B, B , C, C , C , D, E, F, F and G. The variants observed in water buffalo did not exist in the genus. In addition, 14 amino acid differential sites of -CN between buffalo and the genus were identified, of which 3 were located at glycosylation sites (80S, 96T, 141S) and 4 at phosphorylation sites (19S, 80S, 96T, 141S). It is speculated that they may lead to differences in the physicochemical properties of -CN between buffalo and the genus. This study will lay a foundation for exploring the association between the variation in the gene and the lactation traits of buffalo.