Polymorphisms of the kappa casein (CSN3) gene and inference of its variants in water buffalo (Bubalus bubalis)

Kappa casein plays a crucial role in the formation of stable casein micellesand has a key influence on milk-clotting properties. However, currentunderstanding of buffalo CSN3 gene polymorphisms is not sufficient. In thisstudy, the polymorphisms in the complete coding sequence (CDS) of the buffalo CSN3were detected using PCR product direct sequencing. The CDS of CSN3 for river andswamp buffalo was the same in length, which contained an open reading frameof 573 nucleotides encoding a peptide containing 190 amino acid residues. Atotal of eight single nucleotide polymorphisms (SNPs) was identified in twotypes of buffalo. Among them, c.86C>T, c.252G>C,c.445G>A, c.467C>T and c.516A>C werenon-synonymous, which leads to p.Pro8Leu, p.Lys63Asn, p.Val128Ile,p.Thr135Ile and p.Glu151Asp substitutions in buffalo kappa casein (<span class="inline-formula">κ</span>-CN),respectively. The substitution of p.Thr135Ile may exert a vital effect on thefunction of buffalo <span class="inline-formula">κ</span>-CN. Eleven haplotypes were defined based onthe SNPs found in buffalo, and accordingly, seven protein variants and four synonymous variants of buffalo <span class="inline-formula">κ</span>-CN were inferred, called variants A,B, B<span class="inline-formula">¹</span>, C, C<span class="inline-formula">¹</span>, C<span class="inline-formula">²</span>, D, E, F, F<span class="inline-formula">¹</span> and G. Thevariants observed in water buffalo did not exist in the Bos genus. In addition, 14 amino acid differential sites of <span class="inline-formula">κ</span>-CN between buffalo and the Bos genuswere identified, of which 3 were located at glycosylation sites (80S, 96T,141S) and 4 at phosphorylation sites (19S, 80S, 96T, 141S). It is speculatedthat they may lead to differences in the physicochemical properties of<span class="inline-formula">κ</span>-CN between buffalo and the Bos genus. This study will lay a foundationfor exploring the association between the variation in the CSN3 gene and thelactation traits of buffalo.