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We screened 57 Sahiwal cattle (Bos indicus) and 53 Murrah, 19 Nili‐Ravi and 11 Egyptian buffaloes (Bubalus bubalis) to detect the polymorphisms at kappa‐casein (CSN3) gene using the polymerase chain reaction (PCR). CSN3 A and B alleles were identified by PCR‐RFLP using the restriction enzymes that detect the underlying nucleotide changes at codon 136 (TaqI) and at codon 148 (HindIII or HinfI) in cattle. The frequency of CSN3 B allele in the Sahiwal cattle was estimated as 0.16 with no homozygous BB animal. Using the same set of primers as used in the Sahiwal cattle, a part of exon IV of buffalo CSN3 gene was amplified, but restriction enzyme analysis using HindIII/HinfI and TaqI did not reveal any polymorphism. However, DNA sequencing of the amplified fragment (GenBank Acc. No. U96662) revealed one polymorphism at codon 135 (ThrACC ?IleATC) in buffalo; the frequencies of 135 Thr/Ile …